Flea protease proteins, nucleic acid molecules, and uses thereof

ABSTRACT

The present invention relates to flea serine protease proteins and flea cysteine protease proteins; to flea serine protease and cysteine protease nucleic acid molecules, including those that encode such proteins; to antibodies raised against such proteins; and to compounds that inhibit flea serine protease and/or cysteine protease activities. The present invention also includes methods to obtain such proteins, nucleic acid molecules, antibodies, and inhibitors. Also included in the present invention are therapeutic compositions comprising such proteins, nucleic acid molecules, antibodies, and/or inhibitors as well as the use of such therapeutic compositions to protect a host animal from flea infestation.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a divisional of U.S. patent application Ser. No. 08/749,699, filed Nov. 15, 1996, which is a continuation-in-part of U.S. patent application Ser. Nos. 08/484,211 (issued as U.S. Pat. No. 5,972,645 on Oct. 26, 1999), 08/482,130 (issued as U.S. Pat. No. 5,962,257 on Oct. 5, 1999), 08/485,443, and 08/485,455 (now U.S. Pat. No. 5,712,143, issued Jan. 27, 1998), each of which was filed on Jun. 7, 1995 and each of which is a continuation-in-part of U.S. patent application Ser. No. 08/326,773, filed Oct. 18, 1994 now U.S. Pat. No. 5,766,609, issued Jun. 16, 1998, which is a continuation-in-part of U.S. patent application Ser. No. 07/806,482, filed Dec. 13, 1991, which issued as U.S. Pat. No. 5,356,622 on Oct. 18, 1994. The present application is also a continuation-in-part of U.S. patent application Ser. No. 08/326,773, ibid., as well as of WO 96/11706, the international publication date of which is Apr. 25, 1996. WO 96/11706 claims priority to U.S. patent application Ser. Nos. 08/326,773, 08/484,211, 08/482,130, 08/485,443 and 08/485,455, ibid. Each of the applications referred to in this section is incorporated herein by reference in its entirety.

FIELD OF THE INVENTION

The present invention relates to novel flea protease proteins and their use to reduce flea infestation of animals. The present invention also relates to the use of anti-flea protease antibodies and other compounds that reduce flea protease activity to reduce flea infestation of animals.

BACKGROUND OF THE INVENTION

Fleas, which belong to the insect order Siphonaptera, are obligate ectoparasites for a wide variety of animals, including birds and mammals. Flea infestation of animals is of health and economic concern because fleas are known to cause and/or transmit a variety of diseases. Fleas cause and/or carry infectious agents that cause, for example, flea allergy dermatitis, anemia, murine typhus, plague and tapeworm. In addition, fleas are a problem for animals maintained as pets because the infestation becomes a source of annoyance for the pet owner who may find his or her home generally contaminated with fleas which feed on the pets. As such, fleas are a problem not only when they are on an animal but also when they are in the general environment of the animal.

The medical and veterinary importance of flea infestation has prompted the development of reagents capable of controlling flea infestation. Commonly encountered methods to control flea infestation are generally focussed on use of insecticides in formulations such as sprays, shampoos, dusts, dips, or foams, or in pet collars. While some of these products are efficacious, most, at best, offer protection of a very limited duration. Furthermore, many of the methods are often not successful in reducing flea populations on the pet for one or more of the following reasons: (1) failure of owner compliance (frequent administration is required); (2) behavioral or physiological intolerance of the pet to the pesticide product or means of administration; and (3) the emergence of flea populations resistant to the prescribed dose of pesticide. Additional anti-flea products include nontoxic reagents such as insect growth regulators (IGRs), including methoprene, which mimics flea hormones and affect flea larval development.

An alternative method for controlling flea infestation is the use of flea vaccines to be administered to animals prior to or during flea infestation. However, despite considerable interest in developing anti-flea reagents, no flea vaccine presently exists.

SUMMARY OF THE INVENTION

The present invention relates to flea serine protease proteins, to flea aminopeptidase proteins, and to flea cysteine protease proteins; to flea serine protease, aminopeptidase and/or cysteine protease nucleic acid molecules, including those that encode such proteins; to antibodies raised against such proteins; and to compounds that inhibit flea serine protease, aminopeptidase and/or cysteine protease activities. The present invention also includes methods to obtain such proteins, nucleic acid molecules, antibodies, and inhibitors. Also included in the present invention are therapeutic compositions comprising such proteins, nucleic acid molecules, antibodies, and/or inhibitors as well as the use of such therapeutic compositions to protect a host animal from flea infestation.

One embodiment of the present invention is an isolated nucleic acid molecule that hybridizes under stringent hybridization conditions with a gene including a serine protease gene comprising a nucleic acid sequence including a nucleic acid molecule including SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:23, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:28, SEQ ID NO:29, SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:34, SEQ ID NO:35, SEQ ID NO:37, SEQ ID NO:39, SEQ ID NO:.40, SEQ ID NO:42, SEQ ID NO:43 and/or SEQ ID NO:45, and a cysteine protease gene comprising a nucleic acid molecule selected from the group consisting of SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:88, SEQ ID NO:90, SEQ ID NO:91, SEQ ID NO:93 and/or SEQ ID NO:94.

The present invention also includes a nucleic acid molecule that hybridizes under stringent hybridization conditions with a nucleic acid sequence encoding a protein comprising an amino acid sequence including SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:16, SEQ ID NO:19, SEQ ID NO:22, SEQ ID NO:24, SEQ ID NO:27, SEQ ID NO:30, SEQ ID NO:33, SEQ ID NO:36, SEQ ID NO:38, SEQ ID NO:41, SEQ ID NO:44, SEQ ID NO:67, SEQ ID NO:68, SEQ ID NO:69, SEQ ID NO:70, SEQ ID NO:71, SEQ ID NO:72, SEQ ID NO:73, SEQ ID NO:96, SEQ ID NO:2, SEQ ID NO:5, SEQ ID NO:8, SEQ ID NO:89, SEQ ID NO:92 and/or SEQ ID NO:95, or with a nucleic acid sequence that is a complement of any of the nucleic acid sequences. A preferred nucleic acid sequence of the present invention includes a nucleic acid molecule comprising a nucleic acid sequence including SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:23, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:28, SEQ ID NO:29, SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:34, SEQ ID NO:35, SEQ ID NO:37, SEQ ID NO:39, SEQ ID NO:40, SEQ ID NO:42, SEQ ID NO:43 and SEQ ID NO:45, SEQ ID NO:1, SEQ ID NO:3 SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:88, SEQ ID NO:90, SEQ ID NO:91, SEQ ID NO:93 and/or SEQ ID NO:94, and allelic variants thereof.

The present invention also includes an isolated protein encoded by a nucleic acid molecule that hybridizes under stringent hybridization conditions with a nucleic acid molecule having a nucleic acid sequence encoding a protein comprising an amino acid sequence including SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:16, SEQ ID NO:19, SEQ ID NO:22, SEQ ID NO:24, SEQ ID NO:27, SEQ ID NO:30, SEQ ID NO:33, SEQ ID NO:36, SEQ ID NO:38, SEQ ID NO:41, SEQ ID NO:44, SEQ ID NO:67, SEQ ID NO:68, SEQ ID NO:69, SEQ ID NO:70, SEQ ID NO:71, SEQ ID NO:72, SEQ ID NO:73, SEQ ID NO:96, SEQ ID NO:2, SEQ ID NO:5, SEQ ID NO:8, SEQ ID NO:89, SEQ ID NO:92 and SEQ ID NO:95.

The present invention also relates to recombinant molecules, recombinant viruses and recombinant cells that include a nucleic acid molecule of the present invention. Also included are methods to produce such nucleic acid molecules, recombinant molecules, recombinant viruses and recombinant cells.

Yet another embodiment of the present invention is a therapeutic composition that is capable of reducing hematophagous ectoparasite infestation. Such a therapeutic composition includes an excipient and a protective compound including: an isolated protein or mimetope thereof encoded by a nucleic acid molecule that hybridizes under stringent hybridization conditions with a nucleic acid molecule having a nucleic acid sequence encoding a protein comprising an amino acid sequence including SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:16, SEQ ID NO:19, SEQ ID NO:22, SEQ ID NO:24, SEQ ID NO:27, SEQ ID NO:30, SEQ ID NO:33, SEQ ID NO:36, SEQ ID NO:38, SEQ ID NO:41, SEQ ID NO:44, SEQ ID NO:67, SEQ ID NO:68, SEQ ID NO:69, SEQ ID NO:70, SEQ ID NO:71, SEQ ID NO:72, SEQ ID NO:73, SEQ ID NO:96, SEQ ID NO:2, SEQ ID NO:5, SEQ ID NO:8, SEQ ID NO:89, SEQ ID NO:92 and SEQ ID NO:95; an isolated nucleic acid molecule that hybridizes under stringent hybridization conditions with a gene comprising a nucleic acid sequence including SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:23, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:28, SEQ ID NO:29, SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:34, SEQ ID NO:35, SEQ ID NO:37, SEQ ID NO:39, SEQ ID NO:40, SEQ ID NO:42, SEQ ID NO:43, SEQ ID NO:45, SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:88, SEQ ID NO:90, SEQ ID NO:91, SEQ ID NO:93 and SEQ ID NO:94; an isolated antibody that selectively binds to a protein encoded by a nucleic acid molecule that hybridizes under stringent hybridization conditions with a nucleic acid molecule having a nucleic acid sequence encoding a protein comprising an amino acid sequence including SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:16, SEQ ID NO:19, SEQ ID NO:22, SEQ ID NO:24, SEQ ID NO:27, SEQ ID NO:30, SEQ ID NO:33, SEQ ID NO:36, SEQ ID NO:38, SEQ ID NO:41, SEQ ID NO:44, SEQ ID NO:67, SEQ ID NO:68, SEQ ID NO:69, SEQ ID NO:70, SEQ ID NO:71, SEQ ID NO:72, SEQ ID NO:73, SEQ ID NO:96, SEQ ID NO:2, SEQ ID NO:5, SEQ ID NO:8, SEQ ID NO:89, SEQ ID NO:92 and SEQ ID NO:95; an inhibitor of protease activity identified by its ability to inhibit the activity of a protein encoded by a nucleic acid molecule that hybridizes under stringent hybridization conditions with a nucleic acid molecule having a nucleic acid sequence encoding a protein comprising an amino acid sequence including SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:16, SEQ ID NO:19, SEQ ID NO:22, SEQ ID NO:24, SEQ ID NO:27, SEQ ID NO:30, SEQ ID NO:33, SEQ ID NO:36, SEQ ID NO:38, SEQ ID NO:41, SEQ ID NO:44, SEQ ID NO:67, SEQ ID NO:68, SEQ ID NO:69, SEQ ID NO:70, SEQ ID NO:71, SEQ ID NO:72, SEQ ID NO:73, SEQ ID NO:96, SEQ ID NO:2, SEQ ID NO:5, SEQ ID NO:8, SEQ ID NO:89, SEQ ID NO:92 and SEQ ID NO:95; and a mixture thereof. Also included in the present invention is a method to reduce flea infestation, comprising the step of administering to the animal a therapeutic composition of the present invention.

Another embodiment of the present invention is a method to identify a compound capable of inhibiting flea protease activity, the method comprising: (a) contacting an isolated flea protease protein comprising an amino acid sequence including SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:16, SEQ ID NO:19, SEQ ID NO:22, SEQ ID NO:24, SEQ ID NO:27, SEQ ID NO:30, SEQ ID NO:33, SEQ ID NO:36, SEQ ID NO:38, SEQ ID NO:41, SEQ ID NO:44, SEQ ID NO:67, SEQ ID NO:68, SEQ ID NO:69, SEQ ID NO:70, SEQ ID NO:71, SEQ ID NO:72, SEQ ID NO:73, SEQ ID NO:96, SEQ ID NO:2, SEQ ID NO:5, SEQ ID NO:8, SEQ ID NO:89, SEQ ID NO:92 and SEQ ID NO:95 with a putative inhibitory compound under conditions in which, in the absence of said compound, the protein has proteolytic activity; and (b) determining if the putative inhibitory compound inhibits the activity. The present invention also includes a kit to to identify a compound capable of inhibiting flea protease activity.

The present invention also includes an isolated flea protease protein that cleaves an immunoglobulin, when the protein is incubated in the presence of the immunoglobulin in about 100 microliters of about 0.2M Tris-HCl for about 18 hours at about 37° C. A preferred protease protein capable of cleaving immunoglobulin comprises an amino acid sequence selected from the group consisting of SEQ ID NO:67, SEQ ID NO:68, SEQ ID NO:69, SEQ ID NO:70, SEQ ID NO:71, SEQ ID NO:72, SEQ ID NO:73 and SEQ ID NO:96.

Another embodiment of the present invention includes a method to identify a compound capable of inhibiting flea immunoglobulin proteinase protein activity, the method comprising: (a) contacting an isolated flea immunoglobulin proteinase protein with a putative inhibitory compound under conditions in which, in the absence of the compound, the protein has immunoglobulin proteinase activity; and (b) determining if the putative inhibitory compound inhibits the activity.

DETAILED DESCRIPTION OF THE INVENTION

The present invention includes the use of compounds that inhibit flea protease activity to protect a host animal from flea infestation. The inventors have discovered that proteases are significant components of the flea midgut and are good targets for immunotherapeutic and/or chemotherapeutic intervention to reduce flea burden both on the host animal and in the immediate (i.e., surrounding) environment of the animal. The inventors have shown, for example, that the viability and/or fecundity of fleas consuming a blood meal is reduced when the blood meal contains compounds that reduce flea protease activity, probably because the compounds interfere with flea digestion and other functions. Compounds that reduce the amount and/or activity of flea proteases without substantially harming the host animal are included in the present invention. Such compounds include flea protease vaccines, anti-flea protease antibodies, flea protease inhibitors, and/or compounds that suppress protease synthesis; such compounds are discussed in more detail below.

One embodiment of the present invention is a method to protect a host animal from flea infestation by treating the animal with a composition that includes a compound that reduces the protease activity of fleas feeding (includes fleas in the process of feeding as well as fleas having fed) from the treated animal thereby reducing the flea burden on the animal and in the environment of the animal. It is to be noted that the term "a" or "an" entity refers to one or more of that entity; for example, a compound refers to one or more compounds. As such, the terms "a" (or "an"), "one or more" and "at least one" can be used interchangeably herein. Thus, a composition of the present invention can include one or more compounds that target (reduced the activity of) one or more proteases in the flea.

As used herein, the phrase "to protect an animal from flea infestation" refers to reducing the potential for flea population expansion on and around the animal (i.e., reducing the flea burden). Preferably, the flea population size is decreased, optimally to an extent that the animal is no longer bothered by fleas. A host animal, as used herein, is an animal from which fleas can feed by attaching to and feeding through the skin of the animal. Fleas, and other ectoparasites, can live on a host animal for an extended period of time or can attach temporarily to an animal in order to feed. At any given time, a certain percentage of a flea population can be on a host animal whereas the remainder can be in the environment surrounding the animal (i.e., in the environment of the animal). Such an environment can include not only adult fleas, but also flea eggs and/or flea larvae. The environment can be of any size such that fleas in the environment are able to jump onto and off of a host animal. As such, it is desirable not only to reduce the flea burden on an animal per se, but also to reduce the flea burden in the environment surrounding the animal.

In accordance with the present invention, a host animal is treated by administering to the animal a compound of the present invention in such a manner that the compound itself (e.g., a protease inhibitor, protease synthesis suppressor or anti-flea protease antibody) or a product generated by the animal in response to administration of the compound (e.g., antibodies produced in response to a flea protease vaccine, or conversion of an inactive inhibitor "prodrug" to an active protease inhibitor) ultimately enters the flea midgut. An animal is preferably treated in such a way that the compound or product thereof enters the blood stream of the animal. Fleas are then exposed to the compound when they feed from the animal. For example, flea protease inhibitors administered to an animal are administered in such a way that the inhibitors enter the blood stream of the animal, where they can be taken up by feeding fleas. In another embodiment, when a host animal is administered a flea protease vaccine, the treated animal mounts an immune response resulting in the production of antibodies against the protease (anti-flea protease antibodies) which circulate in the animal's blood stream and are taken up by fleas upon feeding. Blood taken up by fleas enters the flea midgut where compounds of the present invention, or products thereof, such as anti-flea protease antibodies, flea protease inhibitors, and/or protease synthesis suppressors, interact with, and reduce proteolytic activity in the flea midgut. The present invention also includes the ability to reduce larval flea infestation in that when fleas feed from a host animal that has been administered a therapeutic composition of the present invention, at least a portion of compounds of the present invention, or products thereof, in the blood taken up by the flea are excreted by the flea in feces, which is subsequently ingested by flea larvae. It is of note that flea larvae obtain most, if not all, of their nutrition from flea feces.

In accordance with the present invention, reducing proteolytic activity in flea midguts can lead to a number of outcomes that reduce flea burden on treated animals and their surrounding environments. Such outcomes include, but are not limited to, (a) reducing the viability of fleas that feed from the treated animal, (b) reducing the fecundity of female fleas that feed from the treated animal, (c) reducing the reproductive capacity of male fleas that feed from the treated animal, (d) reducing the viability of eggs laid by female fleas that feed from the treated animal, (e) altering the blood feeding behavior of fleas that feed from the treated animal (e.g., fleas take up less volume per feeding or feed less frequently), (f) reducing the viability of flea larvae, for example due to the feeding of larvae from feces of fleas that feed from the treated animal and/or (g) altering the development of flea larvae (e.g., by decreasing feeding behavior, inhibiting growth, inhibiting (e.g., slowing or blocking) molting, and/or otherwise inhibiting maturation to adults).

One embodiment of the present invention is a composition that includes one or more compounds that reduce the activity of one or more flea proteases directly (e.g., an anti-flea protease antibody or a flea protease inhibitor) and/or indirectly (e.g., a flea protease vaccine). Suitable flea proteases to target include flea aminopeptidases, flea carboxypeptidases and/or flea endopeptidases. Such proteases can include cytosolic and/or membrane-bound forms of a protease. Preferred flea proteases to target include, but are not limited to, serine proteases, metalloproteases, aspartic acid proteases and/or cysteine proteases. It is to be noted that these preferred groups of proteases include aminopeptidases, carboxypeptidases and/or endopeptidases. Preferred flea proteases to target include, but are not limited to, proteases that degrade hemoglobin, proteases involved in blood coagulation and/or lytic (anti-coagulation) pathways, proteases involved in the maturation of peptide hormones, proteases that inhibit complement or other host immune response elements (e.g., antibodies) and/or proteases involved in vitellogenesis. A number of proteases are known to those skilled in the art, including, but not limited to, aminopeptidases, such as leucine aminopeptidase and aminopeptidases B and M; astacin-like metalloproteases; calpains; carboxypeptidases, such as carboxypeptidases A, P and Y; cathepsins, such as cathepsins B, D, E, G, H, and L, chymotrypsins; cruzipains; meprins; papains; pepsins; renins; thermolysins and trypsins. A particularly preferred protease to target is a protease having a proteolytic activity that, when targeted with a composition of the present invention, reduces flea burden without substantially harming the host animal. Such a protease can be identified using, for example, methods as disclosed herein.

One aspect of the present invention is the discovery that a substantial amount of the proteolytic activity found in flea midguts is serine protease activity. Both in vitro and in vivo studies using a number of protease inhibitors substantiate this discovery, details of which are disclosed in the Examples. As such a particularly preferred protease to target is a serine protease. Examples of serine proteases, include, but are not limited to, acrosins, bromelains, cathepsin G, chymotrypsins, collagenases, elastases, factor Xa, ficins, kallikreins, papains, plasmins, Staphylococcal V8 proteases, thrombins and trypsins. In one embodiment, a preferred flea serine protease to target includes a protease having trypsin-like or chymotrypsin-like activity. It is appreciated by those skilled in the art that an enzyme having "like" proteolytic activity has similar activity to the referenced protease, although the exact structure of the preferred substrate cleaved may differ. "Like" proteases usually have similar tertiary structures as their referenced counterparts.

Protease inhibitor studies disclosed in the Examples section also indicate that additional preferred proteases to target include aminopeptidases and/or metalloproteases. Examples of such proteases include exo- and endo-metalloproteases, digestive enzymes, and enzymes involved in peptide hormone maturation. One example of an aminopeptidase that is also a metalloprotease is leucine aminopeptidase.

Suitable compounds to include in compositions of the present invention include, but are not limited to, a vaccine comprising a flea protease (a flea protease vaccine), an antibody that selectively binds to a flea protease (an anti-flea protease antibody), a flea protease inhibitor (a compound other than a vaccine or an antibody that inhibits a flea protease), and a mixture of such compounds. As used herein, a mixture thereof refers to a combination of one or more of the cited entities. Compositions of the present invention can also include compounds to suppress protease synthesis or maturation, such as, but not limited to, protease modulating peptides.

A preferred embodiment of the present invention is a flea protease vaccine and its use to reduce the flea population on and around an animal. A flea protease vaccine can include one or more proteins capable of eliciting an immune response against a flea protease and can also include other components. Preferred flea protease vaccines include a flea serine protease, a flea metalloprotease, a flea aspartic acid protease and/or a flea cysteine protease, with flea serine protease, flea metalloprotease and/or flea aminopeptidase vaccines being more preferred. Examples of flea protease vaccines include soluble flea midgut preparations of the present invention as well as one or more isolated proteins of the present invention.

One embodiment of the present invention is a soluble flea midgut preparation. Such a preparation includes primarily components naturally present in the lumen of a flea midgut and, depending on the method of preparation, can also include one or more peripheral midgut membrane proteins. Methods to preferentially include, or exclude, membrane proteins from such a preparation are known to those skilled in the art. The present invention includes the discovery that such a preparation has proteolytic activity, of which a substantial portion is serine protease activity. Preferably at least about 70 percent of the proteolytic activity in a soluble flea midgut soluble preparation is serine protease activity, as can be indicated by the ability to inhibit at least about 70 percent of the proteolytic activity with 4-2-aminoethylbenzenesulfonylfluoride-hydrochloride (AEBSF). Serine protease activity can also be identified using other known inhibitors or substrates. Other preferred inhibitors that can inhibit at least about 70 percent of the proteolytic activity of a soluble flea midgut preparation of the present invention include soybean trypsin inhibitor, 1,3-diisopropylfluorophosphate or leupeptin.

A soluble flea midgut preparation of the present invention includes proteases that range in molecular weight from about 5 kilodaltons (kD or kDa) to about 200 kD, as determined by SDS-PAGE (sodium dodecyl sulfate polyacrylamide gel electrophoresis), with at least a substantial portion of the serine proteases ranging in molecular weight from about 5 kD to about 60 kD, as determined by SDS-PAGE. A substantial portion of protease activity in a soluble flea midgut preparation of the present invention has a pH activity optimum ranging from about pH 5 to about pH 10, preferably an activity optimum ranging from about pH 7 to about pH 9, and even more preferably an activity optimum of about pH 8. While not being bound by theory, such a pH optimum suggests that a large proportion of proteases in soluble flea midgut preparations of the present invention are serine proteases. It is also interesting to note that the pH of the flea midgut is also about pH 8. The findings that proteases in soluble flea midgut preparations of the present invention exhibit a varied pattern of inhibition by protease inhibitors of a given type (e.g., serine protease inhibitors), as well as variances seen in molecular weights and pH optima of the proteases, suggest that there are a number of protease isoforms in such preparations.

A soluble flea midgut preparation of the present invention is preferably prepared by a method that includes the steps of (a) disrupting a flea midgut to produce a mixture including a liquid portion and a solid portion and (b) recovering the liquid portion to obtain a soluble flea midgut preparation. Such a method is a simplified version of methods disclosed in U.S. Pat. No. 5,356,622, ibid. It is to be noted that in accordance with the present invention, methods disclosed in U.S. Pat. No. 5,356,622, ibid. can also be used to prepare soluble flea midgut preparations having similar proteolytic activities.

Flea midguts can be obtained (e.g., dissected from) from unfed fleas or from fleas that recently consumed a blood meal (i.e., blood-fed fleas). Such midguts are referred to herein as, respectively, unfed flea midguts and fed flea midguts. Flea midguts can be obtained from either male or female fleas. As demonstrated in the Examples section, female flea midguts exhibit somewhat more proteolytic activity than do male flea midguts. Furthermore, fed flea midguts have significantly more proteolytic activity than do unfed flea midguts. While not being bound by theory, it is believed that blood feeding induces in flea midguts the synthesis and/or activation of proteases as well as other factors (e.g., enzymes, other proteins, co-factors, etc.) important in digesting the blood meal, as well as in neutralizing host molecules potentially damaging to the flea (e.g., complement, immunoglobulins, blood coagulation factors). It is also to be appreciated that unfed flea midguts may contain significant targets not found in fed flea midguts and vice versa. Furthermore, although the present application focuses primarily on flea midgut proteases, it is to be noted that the present invention also includes other components of soluble flea midgut preparations of the present invention that provide suitable targets to reduce flea burden on an animal and in the environment of that animal; see also U.S. Pat. No. 5,356,622, ibid.

Methods to disrupt flea midguts in order to obtain a soluble flea midgut preparation are known to those skilled in the art and can be selected according to, for example, the volume being processed and the buffers being used. Such methods include any technique that promotes cell lysis, such as, but are not limited to, chemical disruption techniques (e.g., exposure of midguts to a detergent) as well as mechanical disruption techniques (e.g., homogenization, sonication, use of a tissue blender or glass beads, and freeze/thaw techniques).

Methods to recover a soluble flea midgut preparation are also known to those skilled in the art and can include any method by which the liquid portion of disrupted flea midguts is separated from the solid portion (e.g., filtration or centrifugation). In a preferred embodiment, disrupted flea midguts are subjected to centrifugation, preferably at an acceleration ranging from about 10,000× g to about 15,000× g for several minutes (e.g., from about 1 min. to about 15 min.). The supernatant from such a centrifugation comprises a soluble flea midgut preparation of the present invention.

The present invention also includes an isolated protein that includes an amino acid sequence encoded by a nucleic acid molecule capable of hybridizing under stringent conditions (i.e., that hybridize under stringent hybridization conditions) with a nucleic acid molecule that encodes a protease present (i.e., the nucleic acid molecules hybridize with the nucleic acid strand that is complementary to the coding strand) in (i.e., can be found in) a flea midgut, such as a midgut from a blood-fed female flea, a midgut from a blood-fed male flea, a midgut from an unfed female flea or a midgut from an unfed male flea. A preferred midgut protease is present in the lumen of the midgut.

An isolated protein of the present invention, also referred to herein as an isolated protease protein, preferably is capable of eliciting an immune response against a flea midgut protease and/or has proteolytic activity. According to the present invention, an isolated, or biologically pure, protein, is a protein that has been removed from its natural milieu. As such, "isolated" and "biologically pure" do not necessarily reflect the extent to which the protein has been purified. An isolated protease protein can be obtained from its natural source. Such an isolated protein can also be produced using recombinant DNA technology or chemical synthesis.

As used herein, an isolated protein of the present invention can be a full-length protein or any homologue of such a protein, such as a protein in which amino acids have been deleted (e.g., a truncated version of the protein, such as a peptide), inserted, inverted, substituted and/or derivatized (e.g., by glycosylation, phosphorylation, acetylation, myristoylation, prenylation, palmitoylation, amidation and/or addition of glycerophosphatidyl inositol) such that the homologue comprises a protein having an amino acid sequence that is sufficiently similar to a natural flea midgut protease that a nucleic acid sequence encoding the homologue is capable of hybridizing under stringent conditions to (i.e., with) the complement of a nucleic acid sequence encoding the corresponding natural flea midgut protease amino acid sequence. As used herein, stringent hybridization conditions refer to standard hybridization conditions under which nucleic acid molecules, including oligonucleotides, are used to identify similar nucleic acid molecules. Such standard conditions are disclosed, for example, in Sambrook et al., Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Labs Press, 1989; Sambrook et al., ibid., is incorporated by reference herein in its entirety. Stringent hybridization conditions typically permit isolation of nucleic acid molecules having at least about 70% nucleic acid sequence identity with the nucleic acid molecule being used to probe in the hybridization reaction. Formulae to calculate the appropriate hybridization and wash conditions to achieve hybridization permitting 30% or less mismatch of nucleotides are disclosed, for example, in Meinkoth et al., 1984, Anal. Biochem. 138, 267-284; Meinkoth et al., ibid., is incorporated by reference herein in its entirety.

The minimal size of a protein homologue of the present invention is a size sufficient to be encoded by a nucleic acid molecule capable of forming a stable hybrid with the complementary sequence of a nucleic acid molecule encoding the corresponding natural protein. As such, the size of the nucleic acid molecule encoding such a protein homologue is dependent on nucleic acid composition and percent homology between the nucleic acid molecule and complementary sequence as well as upon hybridization conditions per se (e.g., temperature, salt concentration, and formamide concentration). The minimal size of such nucleic acid molecules is typically at least about 12 to about 15 nucleotides in length if the nucleic acid molecules are GC-rich and at least about 15 to about 17 bases in length if they are AT-rich. As such, the minimal size of a nucleic acid molecule used to encode a protease protein homologue of the present invention is from about 12 to about 18 nucleotides in length. There is no limit, other than a practical limit, on the maximal size of such a nucleic acid molecule in that the nucleic acid molecule can include a portion of a gene, an entire gene, or multiple genes, or portions thereof. Similarly, the minimal size of a protease protein homologue of the present invention is from about 4 to about 6 amino acids in length, with preferred sizes depending on whether a full-length, multivalent (i.e., fusion protein having more than one domain each of which has a function), or functional portions of such proteins are desired. Protease protein homologues of the present invention preferably have protease activity and/or are capable of eliciting an immune response against a flea midgut protease.

A protease protein homologue of the present invention can be the result of allelic variation of a natural gene encoding a flea protease. A natural gene refers to the form of the gene found most often in nature. Protease protein homologues can be produced using techniques known in the art including, but not limited to, direct modifications to a gene encoding a protein using, for example, classic or recombinant DNA techniques to effect random or targeted mutagenesis. Isolated protease proteins of the present invention, including homologues, can be identified in a straight-forward manner by the proteins' ability to effect proteolytic activity and/or to elicit an immune response against a flea midgut protease. Such techniques are known to those skilled in the art.

A preferred protease protein of the present invention is a flea serine protease, a flea metalloprotease, a flea aspartic acid protease, a flea cysteine protease, or a homologue of any of these proteases. A more preferred protease protein is a flea serine protease, a flea metalloprotease or a homologue of either. Also preferred is a flea aminopeptidase or a homologue thereof. Also preferred is a flea cysteine protease or a homologue thereof. Particularly preferred is a flea serine protease or a homologue thereof.

Preferred protease proteins of the present invention are flea protease proteins having molecular weights ranging from about 5 kD to about 200 kD, as determined by SDS-PAGE, and homologues of such proteins. More preferred are flea protease proteins having molecular weights ranging from about 5 kD to about 60 kD, as determined by SDS-PAGE, and homologues of such proteins. Even more preferred are flea serine protease proteins, particularly those having molecular weights of about 26 kD (denoted PfSP26, now denoted PafSP-26K to distinguish from flea PfSP26 as described in Example 26), about 24 kD (denoted PfSP24, now denoted PafSP-24K to distinguish from flea PfSP24 as described in Example 27), about 19 kD (denoted PfSP19, now denoted PafSP-19K to distinguish from flea PfSP19 as described in Example 32), about 6 kD (denoted PfSP6, now denoted PafSP-6K to distinguish from flea PfSP6 as described in Example 11), about 31 kD (denoted PfSP28), about 25 kD (denoted PlfSP-25K1) from 1st instar larvae, about 25 kD (denoted PlfSP-25K3) from 3rd instar larvae, about 28 kD (denoted PlfSP-28K3) and about 31 kD (denoted PlfSP-31K3), and flea aminopeptidase proteins, particularly those having molecular weights of about 95 kD (denoted PfAP-95K) as determined by SDS-PAGE, and homologues of such proteins.

One preferred embodiment of the present invention is an isolated flea protease protein that includes an amino acid sequence encoded by a nucleic acid molecule that hybridizes under stringent hybridization conditions with a flea serine protease gene, with a flea aminopeptidase gene or with a flea cysteine protease gene. As used herein, a flea protease gene includes all nucleic acid sequences related to a natural flea protease gene such as regulatory regions that control production of a flea protease protein encoded by that gene (such as, but not limited to, transcription, translation or post-translation control regions) as well as the coding region itself.

The inventors have discovered an extensive family of serine proteases, encoded by a family of serine protease genes. Such a gene family may be due to allelic variants (i.e., genes having similar, but different, sequences at a given locus in a population of fleas) and/or to, the existence of serine protease genes at more than one locus in the flea genome. As such, the present invention includes flea serine protease genes comprising not only the nucleic acid sequences disclosed herein (e.g., genes including nucleic acid sequences SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:23, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:28, SEQ ID NO:29, SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:34, SEQ ID NO:35, SEQ ID NO:37, SEQ ID NO:39, SEQ ID NO:40, SEQ ID NO:42, SEQ ID NO:43, SEQ ID NO:45, and/or nucleic acid sequences encoding proteins having amino acid sequences as disclosed herein (e.g., SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:16, SEQ ID NO:19, SEQ ID NO:22, SEQ ID NO:24, SEQ ID NO:27, SEQ ID NO:30, SEQ ID NO:33, SEQ ID NO:36, SEQ ID NO:38, SEQ ID NO:41, SEQ ID NO:44, SEQ ID NO:67, SEQ ID NO:68, SEQ ID NO:69, SEQ ID NO:70, SEQ ID NO:71, SEQ ID NO:72, SEQ ID NO:73 and/or SEQ ID NO:96, but also allelic variants of any of those nucleic acid sequences, as well as other nucleic acid molecules and amino acid sequences disclosed in the examples section. (It should be noted that since nucleic acid sequencing technology is not entirely error-free, all sequences represented herein are at best apparent (i.e., deduced) nucleic acid or amino acid sequences.)

A preferred flea cysteine protease gene includes nucleic acid sequence SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:88, SEQ ID NO:90, SEQ ID NO:91, SEQ ID NO:93 and/or SEQ ID NO:94, which encode a cysteine protease protein including SEQ ID NO:2, SEQ ID NO:5, SEQ ID NO:8, SEQ ID NO:89, SEQ ID NO:92 or SEQ ID NO:95. Additional preferred cysteine protease genes include allelic variants of SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:88, SEQ ID NO:90, SEQ ID NO:91, SEQ ID NO:93 and/or SEQ ID NO:94.

A preferred flea serine protease protein of the present invention is encoded by a nucleic acid molecule that hybridizes under stringent hybridization conditions with at least one of the following nucleic acid molecules: nfSP18, nfSP24, nfSP28, nfSP32, nfSP33 and nfSP40. As used herein, each of these nucleic acid molecules represent the entire coding region of a flea serine protease gene of the present invention (at least portions of which are also referred to by flea clone numbers, as described in the Examples). Nucleic acid molecules that contain partial coding regions or other parts of the corresponding gene are denoted by names that include the size of those nucleic acid molecules (e.g., nfSP40₄₂₈). Nucleic acid molecules containing apparent full length coding regions for which the size is known also are denoted by names that include the size of those nucleic acid molecules (e.g., nfSP40₈₄₁). The production, and at least partial nucleic acid sequence, of such nucleic acid molecules is disclosed in the Examples.

Particularly preferred serine protease proteins are encoded by a nucleic acid molecule that hybridizes under stringent hybridization conditions with at least one of the following nucleic acid molecules: nfSP18₅₃₄, nfSP18₇₇₅, nfSP18₂₂₅, nfSP24₄₁₀, nfSP24₁₀₈₉, nfSP24₇₇₄, nfSP24₇₁₁, nfSP28₃₂₃, nfSP32₉₃₃, nfSP32₉₃₃, nfSP32₃₂₄, nfSP32₆₉₉, nfSP33₄₂₆, nfSP33₇₇₈, nfSP33₁₈₉₄, nfSP33₁₂₀₀, nfSP33₇₂₆, nfSP40₈₄₁ and/or nfSP40₇₁₇. Even more preferred serine protease proteins include the following amino acid sequences: SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:16, SEQ ID NO:19, SEQ ID NO:22, SEQ ID NO:24, SEQ ID NO:27, SEQ ID NO:30, SEQ ID NO:33, SEQ ID NO:36, SEQ ID NO:38, SEQ ID NO:41, SEQ ID NO:44, SEQ ID NO:67, SEQ ID NO:68, SEQ ID NO:69, SEQ ID NO:70, SEQ ID NO:71, SEQ ID NO:72, SEQ ID NO:73, SEQ ID NO:89, SEQ ID NO:92, SEQ ID NO:95 and/or SEQ ID No:96. Additional particularly preferred serine protease proteins are encoded by allelic variants of nucleic acid molecules encoding proteins that include the cited amino acid sequences. Also preferred are flea serine protease proteins including regions that have at least about 50%, preferably at least about 75%, and more preferably at least about 90% identity with flea serine protease proteins having amino acid sequences as cited herein.

One embodiment of the present invention is a flea serine protease that degrades immunoglobulin circulating in a host animal (i.e., flea immunoglobulin proteinase or IgGase). An example of a flea immunoglobulin proteinase is presented in the Examples section. Preferably, an immunoglobulin proteinase of the present invention cleaves an immunoglobulin when the protein is incubated in the presence of the immunoglobulin in about 100 microliters of about 0.2M Tris-HCl for about 18 hours at about 37° C. Suitable immunoglobulin proteinase proteins of the present invention are capable of cleaving the hinge region of an immunoglobulin heavy chain. The hinge region of an immunoglobulin is the flexible domain that joins the Fab arms of the immunoglobulin to the Fc portion of the molecule. A more preferred immunoglobulin proteinase protein includes a protein having a molecular weight ranging from about 25 kD to about 35 kD and more preferably having a molecular weight of about 31 kD, in its mature form. An even more preferred immunoglobulin proteinase protein includes a protein comprising an amino acid sequence including SEQ ID NO:67, SEQ ID NO:68, SEQ ID NO:69, SEQ ID NO:70, SEQ ID NO:71, SEQ ID NO:72, SEQ ID NO:73 and/or SEQ ID NO:96, which can be encoded by a gene comprising nucleic acid sequence SEQ ID NO:66. Without being bound by theory, the proteinase activity of an immunoglobulin proteinase of the present invention cleaves an immunoglobulin in such a manner that the immunoglobulin maintains intact heavy and light chain pairs, either as two Fab fragments or one F(ab')₂ fragment. As used herein, a Fab fragment refers to complete immunoglobulin light chains paired with the variable region and CH1 domains of an immunoglobulin heavy chain. As used herein, a F(ab')₂ fragment refers to two Fab fragments that remain linked by a disulfide bond. Both Fab and F(ab')₂ fragments are capable of binding antigen.

A preferred flea cysteine protease protein of the present invention is encoded by a nucleic acid molecule that hybridizes under stringent hybridization conditions with nucleic acid molecule nfCP1 (a flea cysteine protease full-length coding region that includes nfCP1₅₇₃ or nfCP1₁₀₉ (the production of which are described in the Examples). Even more preferred is a cysteine protease that includes amino acid sequence SEQ ID NO:2, SEQ ID NO:5, SEQ ID NO: 8, SEQ ID NO: 89, SEQ ID NO:92, SEQ ID NO:95, or a cysteine protease encoded by an allelic variant of a nucleic acid molecule that includes SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:88, SEQ ID NO:90, SEQ ID NO:91, SEQ ID NO:93 or SEQ ID NO:94. Also preferred is a flea cysteine protease protein including regions that have at least about 50%, preferably at least about 75%, and more preferably at least about 90% identity with SEQ ID NO: 2, SEQ ID NO: 5, SEQ ID NO: 8, SEQ ID NO:89, SEQ ID NO:92 or SEQ ID NO:95.

One embodiment of the present invention is an isolated protein having proteolytic activity that is substantially inhibited by a serine protease inhibitor, an aminopeptidase inhibitor and/or a cysteine protease inhibitor. Such inhibition can be measured by techniques known to those skilled in the art. To be substantially inhibited means, for example, for a serine protease, that at least half of the proteolytic activity of the protease protein is inhibited by a serine protease inhibitor. Preferably at least about 70 percent, and even more preferably at least about 90 percent of the proteolytic activity of the protease protein is inhibited by a serine protease inhibitor. Preferred serine protease inhibitors include flea serpin proteins, and peptides or analogs thereof.

An isolated protein of the present invention can be produced in a variety of ways, including recovering such a protein from a flea midgut and producing such a protein recombinantly. In one embodiment, a flea midgut protease can be recovered by methods heretofore disclosed for obtaining a soluble flea midgut preparation. A flea midgut protease protein can be further purified from a disrupted flea midgut by a number of techniques known to those skilled in the art, including, but not limited to, affinity chromatography, ion exchange chromatography, filtration, electrophoresis (e.g., standard, capillary and flow-through electrophoresis), hydrophobic interaction chromatography, gel filtration chromatography, reverse phase chromatography, concanavalin A chromatography, chromatofocusing and differential solubilization. In one embodiment, a flea midgut protease is purified using protease inhibitor affinity chromatography, an example of which is disclosed in the Examples section.

Another embodiment of the present invention is a method to produce an isolated protein of the present invention using recombinant DNA technology. Such a method includes the steps of (a) culturing a recombinant cell comprising a nucleic acid molecule encoding a protein of the present invention to produce the protein and (b) recovering the protein therefrom. Details on producing recombinant cells and culturing thereof are presented below. The phrase "recovering the protein" refers simply to collecting the whole fermentation medium containing the protein and need not imply additional steps of separation or purification. Proteins of the present invention can be purified using a variety of standard protein purification techniques, as heretofore disclosed.

Isolated proteins of the present invention are preferably retrieved in "substantially pure" form. As used herein, "substantially pure" refers to a purity that allows for the effective use of the protein as a vaccine. A vaccine for animals, for example, should exhibit no substantial toxicity and should be capable of stimulating the production of antibodies in a vaccinated animal.

Another embodiment of the present invention is an isolated nucleic acid molecule capable of hybridizing under stringent conditions with a gene encoding a flea protease present in a flea midgut. Such a nucleic acid molecule is also referred to herein as a flea protease nucleic acid molecule. Particularly preferred is an isolated nucleic acid molecule that hybridizes under stringent conditions with a flea serine protease gene, with a flea aminopeptidase gene or with a flea cysteine protease gene. The characteristics of such genes are disclosed herein. In accordance with the present invention, an isolated nucleic acid molecule is a nucleic acid molecule that has been removed from its natural milieu (i.e., that has been subject to human manipulation). As such, "isolated" does not reflect the extent to which the nucleic acid molecule has been purified. An isolated nucleic acid molecule can include DNA, RNA, or derivatives of either DNA or RNA.

As stated above, a flea protease gene includes all nucleic acid sequences related to a natural flea protease gene such as regulatory regions that control production of a flea protease protein encoded by that gene (such as, but not limited to, transcription, translation or post-translation control regions) as well as the coding region itself. A nucleic acid molecule of the present invention can be an isolated natural flea protease nucleic acid molecule or a homologue thereof. A nucleic acid molecule of the present invention can include one or more regulatory regions, full-length or partial coding regions, or combinations thereof. The minimal size of a flea protease nucleic acid molecule of the present invention is the minimal size capable of forming a stable hybrid under stringent hybridization conditions with a corresponding natural gene. Flea protease nucleic acid molecules can also include a nucleic acid molecule encoding a hybrid protein, a fusion protein, a multivalent protein or a truncation fragment.

An isolated nucleic acid molecule of the present invention can be obtained from its natural source either as an entire (i.e., complete) gene or a portion thereof capable of forming a stable hybrid with that gene. As used herein, the phrase "at least a portion of" an entity refers to an amount of the entity that is at least sufficient to have the functional aspects of that entity. For example, at least a portion of a nucleic acid sequence, as used herein, is an amount of a nucleic acid sequence capable of forming a stable hybrid with the corresponding gene under stringent hybridization conditions.

An isolated nucleic acid molecule of the present invention can also be produced using recombinant DNA technology (e.g., polymerase chain reaction (PCR) amplification, cloning) or chemical synthesis. Isolated flea protease nucleic acid molecules include natural nucleic acid molecules and homologues thereof, including, but not limited to, natural allelic variants and modified nucleic acid molecules in which nucleotides have been inserted, deleted, substituted, and/or inverted in such a manner that such modifications do not substantially interfere with the nucleic acid molecule's ability to encode a flea protease protein of the present invention or to form stable hybrids under stringent conditions with natural nucleic acid molecule isolates.

A flea protease nucleic acid molecule homologue can be produced using a number of methods known to those skilled in the art (see, for example, Sambrook et al., ibid.). For example, nucleic acid molecules can be modified using a variety of techniques including, but not limited to, classic mutagenesis techniques and recombinant DNA techniques, such as site-directed mutagenesis, chemical treatment of a nucleic acid molecule to induce mutations, restriction enzyme cleavage of a nucleic acid fragment, ligation of nucleic acid fragments, polymerase chain reaction (PCR) amplification and/or mutagenesis of selected regions of a nucleic acid sequence, synthesis of oligonucleotide mixtures and ligation of mixture groups to "build" a mixture of nucleic acid molecules and combinations thereof. Nucleic acid molecule homologues can be selected from a mixture of modified nucleic acids by screening for the function of the protein encoded by the nucleic acid (e.g., the ability of a homologue to elicit an immune response against a flea protease and/or to have proteolytic activity) and/or by hybridization with isolated flea protease nucleic acids under stringent conditions.

An isolated flea protease nucleic acid molecule of the present invention can include a nucleic acid sequence that encodes at least one flea protease protein of the present invention, examples of such proteins being disclosed herein. Although the phrase "nucleic acid molecule" primarily refers to the physical nucleic acid molecule and the phrase "nucleic acid sequence" primarily refers to the sequence of nucleotides on the nucleic acid molecule, the two phrases can be used interchangeably, especially with respect to a nucleic acid molecule, or a nucleic acid sequence, being capable of encoding an flea protease protein.

One embodiment of the present invention is a flea protease nucleic acid molecule of the present invention that is capable of hybridizing under stringent conditions to a nucleic acid strand that encodes at least a portion of a flea protease or a homologue thereof or to the complement of such a nucleic acid strand. A nucleic acid sequence complement of any nucleic acid sequence of the present invention refers to the nucleic acid sequence of the nucleic acid strand that is complementary to (i.e., can form a complete double helix with) the strand for which the sequence is cited. It is to be noted that a double-stranded nucleic acid molecule of the present invention for which a nucleic acid sequence has been determined for one strand, that is represented by a SEQ ID NO, also comprises a complementary strand having a sequence that is a complement of that SEQ ID NO. As such, nucleic acid molecules of the present invention, which can be either double-stranded or single-stranded, include those nucleic acid molecules that form stable hybrids under stringent hybridization conditions with either a given SEQ ID NO denoted herein and/or with the complement of that SEQ ID NO, which may or may not be denoted herein. Methods to deduce a complementary sequence are known to those skilled in the art. Preferred is a flea protease nucleic acid molecule that includes a nucleic acid sequence having at least about 65 percent, preferably at least about 75 percent, more preferably at least about 85 percent, and even more preferably at least about 95 percent homology with the corresponding region(s) of the nucleic acid sequence encoding at least a portion of a flea protease protein. Particularly preferred is a flea protease nucleic acid molecule capable of encoding at least a portion of a flea protease that naturally is present in flea midguts and preferably is included in a soluble flea midgut preparation of the present invention. Examples of nucleic acid molecules of the present invention are disclosed in the Examples section.

A preferred flea serine protease nucleic acid molecule of the present invention is a nucleic acid molecule that hybridizes under stringent hybridization conditions with at least one of the following nucleic acid molecules: nfSP18, nfSP24, nfSP28, nfSP32, nfSP33 and/or nfSP40. More preferred is a nucleic acid molecule that hybridizes under stringent hybridization conditions with at least one of the following nucleic acid molecules: nfSP18₅₃₄, nfSP18₇₇₅, nfSP18₂₂₅, nfSP24₄₁₀, nfSP24₁₀₈₉, nfSP24₇₇₄, nfSP24₇₁₁, nfSP28₇₁₁, nfSP28₃₂₃, nfSP32₉₃₃, nfSP32₉₂₄, nfSP32₆₉₉, nfSP33₄₂₆, nfSP33₇₇₈, nfSP33₁₈₉₄, nfSP33₁₂₀₀, nfSP33₇₂₆, nfSP40₈₄₁ and/or nfSP40₇₁₇, as well as other specific nucleic acid molecules disclosed in the Examples section. Even more preferred are nucleic acid molecules that include nfSP18, nfSP24, nfSP28, nfSP32, nfSP33 and/or nfSP40 and even more nfSP18₅₃₄, nfSP18₇₇₅, nfSP18₂₂₅, nfSP24₄₁₀, nfSP24₁₀₈₉, nfSP24₇₇₄, nfSP24₇₁₁, nfSP28₉₂₃, nfSP₃₂ ₉₃₃, nfSP32₉₃₃, nfSP32₉₂₄, nfSP32₆₉₉, nfSP33₄₂₆, nfSP33₇₇₈, nfSP33₁₈₉₄, nfSP33₁₂₀₀, nfSP33₇₂₆, nfSP40₈₄₁ and/or nfSP40₇₁₇, as well as other specific nucleic acid molecules disclosed in the Examples section.

Particularly preferred flea serine protease nucleic acid molecules include at least one of the following sequences: SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:23, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:28, SEQ ID NO:29, SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:34, SEQ ID NO:35, SEQ ID NO:37, SEQ ID NO:39, SEQ ID NO:40, SEQ ID NO:42, SEQ ID NO:43 and/or SEQ ID NO:45, and complements thereof, as well as other specific nucleic acid molecules disclosed in the Examples section. Also preferred are allelic variants of such nucleic acid molecules.

A preferred flea cysteine protease nucleic acid molecule of the present invention is a nucleic acid molecule that hybridizes under stringent hybridization conditions with nfCP1₅₇₃, or nfCP1₁₁₀₉ (the production of which are described in the Examples). More preferred is a cysteine protease nucleic acid molecule that includes nfCP1₅₇₃ or nfCP1₁₁₀₉. Particularly preferred is a nucleic acid molecule that includes nucleic acid sequence SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:88, SEQ ID NO:90, SEQ ID NO:91, SEQ ID NO:93 and/or SEQ ID NO:94, or allelic variants of such nucleic acid molecules.

Knowing a nucleic acid molecule of a flea protease protein of the present invention allows one skilled in the art to make copies of that nucleic acid molecule as well as to obtain a nucleic acid molecule including additional portions of flea protease protein-encoding genes (e.g., nucleic acid molecules that include the translation start site and/or transcription and/or translation control regions), and/or flea protease nucleic acid molecule homologues. Knowing a portion of an amino acid sequence of a flea protease protein of the present invention allows one skilled in the art to clone nucleic acid sequences encoding such a flea protease protein. In addition, a desired flea protease nucleic acid molecule can be obtained in a variety of ways including screening appropriate expression libraries with antibodies which bind to flea protease proteins of the present invention; traditional cloning techniques using oligonucleotide probes of the present invention to screen appropriate libraries or DNA; and PCR amplification of appropriate libraries, or RNA or DNA using oligonucleotide primers of the present invention (genomic and/or cDNA libraries can be used). To isolate flea protease nucleic acid molecules, preferred cDNA libraries include cDNA libraries made from unfed whole fleas, fed whole fleas, fed flea midguts, unfed flea midguts, and flea salivary glands. Techniques to clone and amplify genes are disclosed, for example, in Sambrook et al., ibid. The Examples section includes examples of the isolation of cDNA sequences encoding flea protease proteins of the present invention.

The present invention also includes nucleic acid molecules that are oligonucleotides capable of hybridizing, under stringent conditions, with complementary regions of other, preferably longer, nucleic acid molecules of the present invention that encode at least a portion of a flea protease protein oligonucleotides of the present invention can be RNA, DNA, or derivatives of either. The minimal size of such oligonucleotides is the size required to form a stable hybrid between a given oligonucleotide and the complementary sequence on another nucleic acid molecule of the present invention. Minimal size characteristics are disclosed herein. The size of the oligonucleotide must also be sufficient for the use of the oligonucleotide in accordance with the present invention. Oligonucleotides of the present invention can be used in a variety of applications including, but not limited to, as probes to identify additional nucleic acid molecules, as primers to amplify or extend nucleic acid molecules or in therapeutic applications to inhibit flea protease production. Such therapeutic applications include the use of such oligonucleotides in, for example, antisense-, triplex formation-, ribozyme- and/or RNA drug-based technologies. The present invention, therefore, includes such oligonucleotides and methods to interfere with the production of flea protease proteins by use of one or more of such technologies.

The present invention also includes a recombinant vector, which includes a flea protease nucleic acid molecule of the present invention inserted into any vector capable of delivering the nucleic acid molecule into a host cell. Such a vector contains heterologous nucleic acid sequences, that is nucleic acid sequences that are not naturally found adjacent to flea protease nucleic acid molecules of the present invention. The vector can be either RNA or DNA, either prokaryotic or eukaryotic, and typically is a virus or a plasmid. Recombinant vectors can be used in the cloning, sequencing, and/or otherwise manipulating of flea protease nucleic acid molecules of the present invention. One type of recombinant vector, herein referred to as a recombinant molecule and described in more detail below, can be used in the expression of nucleic acid molecules of the present invention. Preferred recombinant vectors are capable of replicating in the transformed cell. Preferred nucleic acid molecules to include in recombinant vectors of the present invention are disclosed herein.

As heretofore disclosed, one embodiment of the present invention is a method to produce a flea protease protein of the present invention by culturing a cell capable of expressing the protein under conditions effective to produce the protein, and recovering the protein. A preferred cell to culture is a recombinant cell that is capable of expressing the flea protease protein, the recombinant cell being produced by transforming a host cell with one or more nucleic acid molecules of the present invention. Transformation of a nucleic acid molecule into a cell can be accomplished by any method by which a nucleic acid molecule can be inserted into the cell. Transformation techniques include, but are not limited to, transfection, electroporation, microinjection, lipofection, adsorption, and protoplast fusion. A recombinant cell may remain unicellular or may grow into a tissue, organ or a multicellular organism. Transformed nucleic acid molecules of the present invention can remain extrachromosomal or can integrate into one or more sites within a chromosome of the transformed (i.e., recombinant) cell in such a manner that their ability to be expressed is retained. Preferred nucleic acid molecules with which to transform a host cell are disclosed herein.

Suitable host cells to transform include any cell that can be transformed and that can express the introduced flea protease protein. Such cells are, therefore, capable of producing flea protease proteins of the present invention after being transformed with at least one nucleic acid molecule of the present invention. Host cells can be either untransformed cells or cells that are already transformed with at least one nucleic acid molecule. Suitable host cells of the present invention can include bacterial, fungal (including yeast), insect, animal and plant cells. Preferred host cells include bacterial, yeast, insect and mammalian cells, with bacterial (e.g., E. coli) and insect (e.g., Spodoptera) cells being particularly preferred.

A recombinant cell is preferably produced by transforming a host cell with one or more recombinant molecules, each comprising one or more nucleic acid molecules of the present invention operatively linked to an expression vector containing one or more transcription control sequences. The phrase operatively linked refers to insertion of a nucleic acid molecule into an expression vector in a manner such that the molecule is able to be expressed when transformed into a host cell. As used herein, an expression vector is a DNA or RNA vector that is capable of transforming a host cell and of effecting expression of a specified nucleic acid molecule. Preferably, the expression vector is also capable of replicating within the host cell. Expression vectors can be either prokaryotic or eukaryotic, and are typically viruses or plasmids. Expression vectors of the present invention include any vectors that function (i.e., direct gene expression) in recombinant cells of the present invention, including in bacterial, fungal, insect, animal, and/or plant cells. As such, nucleic acid molecules of the present invention can be operatively linked to expression vectors containing regulatory sequences such as promoters, operators, repressors, enhancers, termination sequences, origins of replication, and other regulatory sequences that are compatible with the recombinant cell and that control the expression of nucleic acid molecules of the present invention. As used herein, a transcription control sequence includes a sequence which is capable of controlling the initiation, elongation, and termination of transcription. Particularly important transcription control sequences are those which control transcription initiation, such as promoter, enhancer, operator and repressor sequences. Suitable transcription control sequences include any transcription control sequence that can function in at least one of the recombinant cells of the present invention. A variety of such transcription control sequences are known to those skilled in the art. Preferred transcription control sequences include those which function in bacterial, yeast, helminth, insect and mammalian cells, such as, but not limited to, tac, lac, tzp, trc, oxy-pro, omp/lpp, rrnB, bacteriophage lambda (λ) (such as λp_(L) and λp_(R) and fusions that include such promoters), bacteriophage T7, T7lac, bacteriophage T3, bacteriophage SP6, bacteriophage SPol, metallothionein, alpha mating factor, Pichia alcohol oxidase, alphavirus subgenomic promoters (such as Sindbis virus subgenomic promoters), baculovirus, Heliothis zea insect virus, vaccinia virus, herpesvirus, poxvirus, adenovirus, simian virus 40, retrovirus actin, retroviral long terminal repeat, Rous sarcoma virus, heat shock, phosphate and nitrate transcription control sequences as well as other sequences capable of controlling gene expression in prokaryotic or eukaryotic cells. Additional suitable transcription control sequences include tissue-specific promoters and enhancers as well as lymphokine-inducible promoters (e.g., promoters inducible by interferons or interleukins). Transcription control sequences of the present invention can also include naturally occurring transcription control sequences naturally associated with a DNA sequence encoding a flea protease protein.

Expression vectors of the present invention may also contain secretory signals (i.e., signal segment nucleic acid sequences) to enable an expressed flea protease protein to be secreted from the cell that produces the protein. Suitable signal segments include a flea protease protein signal segment or any heterologous signal segment capable of directing the secretion of a flea protease protein, including fusion proteins, of the present invention. Preferred signal segments include, but are not limited to, flea protease, tissue plasminogen activator (t-PA), interferon, interleukin, growth hormone, histocompatibility and viral envelope glycoprotein signal segments.

Expression vectors of the present invention may also contain fusion sequences which lead to the expression of inserted nucleic acid molecules of the present invention as fusion proteins. Inclusion of a fusion sequence as part of a flea protease nucleic acid molecule of the present invention can enhance the stability during production, storage and/or use of the protein encoded by the nucleic acid molecule. Furthermore, a fusion segment can function as a tool to simplify purification of a flea protease protein, such as to enable purification of the resultant fusion protein using affinity chromatography. A suitable fusion segment can be a domain of any size that has the desired function (e.g., increased stability and/or purification tool). It is within the scope of the present invention to use one or more fusion segments. Fusion segments can be joined to amino and/or carboxyl termini of a flea protease protein. Linkages between fusion segments and flea protease proteins can be constructed to be susceptible to cleavage to enable straight-forward recovery of the flea protease proteins. Fusion proteins are preferably produced by culturing a recombinant cell transformed with a fusion nucleic acid sequence that encodes a protein including the fusion segment attached to either the carboxyl and/or amino terminal end of a flea protease protein.

A recombinant molecule of the present invention is a molecule that can include at least one of any nucleic acid molecule heretofore described operatively linked to at least one of any transcription control sequence capable of effectively regulating expression of the nucleic acid molecule(s) in the cell to be transformed. A preferred recombinant molecule includes one or more nucleic acid molecules of the present invention, with those that encode one or more flea protease proteins, and particularly one or more flea serine protease, aminopeptidase and/or cysteine protease proteins, being more preferred. Similarly, a preferred recombinant cell includes one or more nucleic acid molecules of the present invention, with those that encode one or more flea protease proteins, and particularly one or more flea serine protease, aminopeptidase, and/or cysteine protease proteins, being more preferred.

It may be appreciated by one skilled in the art that use of recombinant DNA technologies can improve expression of transformed nucleic acid molecules by manipulating, for example, the number of copies of the nucleic acid molecules within a host cell, the efficiency with which those nucleic acid molecules are transcribed, the efficiency with which the resultant transcripts are translated, and the efficiency of post-translational modifications. Recombinant techniques useful for increasing the expression of nucleic acid molecules of the present invention include, but are not limited to, operatively linking nucleic acid molecules to high-copy number plasmids, integration of the nucleic acid molecules into one or more host cell chromosomes, addition of vector stability sequences to plasmids, substitutions or modifications of transcription control signals (e.g., promoters, operators, enhancers), substitutions or modifications of translational control signals (e.g., ribosome binding sites, Shine-Dalgarno sequences), modification of nucleic acid molecules of the present invention to correspond to the codon usage of the host cell, deletion of sequences that destabilize transcripts, and use of control signals that temporally separate recombinant cell growth from recombinant protein production during fermentation. The activity of an expressed recombinant protein of the present invention may be improved by fragmenting, modifying, or derivatizing the resultant protein.

In accordance with the present invention, recombinant cells can be used to produce flea protease proteins of the present invention by culturing such cells under conditions effective to produce such a protein, and recovering the protein. Effective conditions to produce a protein include, but are not limited to, appropriate media, bioreactor, temperature, pH and oxygen conditions that permit protein production. An appropriate, or effective, medium refers to any medium in which a cell of the present invention, when cultured, is capable of producing a flea protease protein. Such a medium is typically an aqueous medium comprising assimilable carbohydrate, nitrogen and phosphate sources, as well as appropriate salts, minerals, metals and other nutrients, such as vitamins. The medium may comprise complex nutrients or may be a defined minimal medium.

Cells of the present invention can be cultured in conventional fermentation bioreactors, which include, but are not limited to, batch, fed-batch, cell recycle, and continuous fermentors. Culturing can also be conducted in shake flasks, test tubes, microtiter dishes, and petri plates. Culturing is carried out at a temperature, pH and oxygen content appropriate for the recombinant cell. Such culturing conditions are well within the expertise of one of ordinary skill in the art.

Depending on the vector and host system used for production, resultant flea protease proteins may either remain within the recombinant cell; be secreted into the fermentation medium; be secreted into a space between two cellular membranes, such as the periplasmic space in E. coli; or be retained on the outer surface of a cell or viral membrane. Methods to purify such proteins are heretofore disclosed.

The present invention also includes isolated anti-flea protease antibodies and their use to reduce flea infestation on a host animal as well as in the environment of the animal. An anti-flea protease antibody is an antibody capable of selectively binding to a protease present in a flea midgut, including female and male fed midguts as well as female and male unfed midguts. An anti-flea protease antibody preferably binds to the protease in such a way as to reduce the proteolytic activity of that protease.

Isolated antibodies are antibodies that have been removed from their natural milieu. The term "isolated" does not refer to the state of purity of such antibodies. As such, isolated antibodies can include anti-sera containing such antibodies, or antibodies that have been purified to varying degrees. As used herein, the term "selectively binds to" refers to the ability of such antibodies to preferentially bind to the protease against which the antibody was raised (i.e., to be able to distinguish that protease from unrelated components in a mixture.). Binding affinities typically range from about 10³ M⁻¹ to about 10¹² M¹. Binding can be measured using a variety of methods known to those skilled in the art including immunoblot assays, immunoprecipitation assays, radioimmunoassays, enzyme immunoassays (e.g., ELISA), immunofluorescent antibody assays and immunoelectron microscopy; see, for example, Sambrook et al., ibid.

Antibodies of the present invention can be either polyclonal or monoclonal antibodies. Antibodies of the present invention include functional equivalents such as antibody fragments and genetically-engineered antibodies, including single chain antibodies, that are capable of selectively binding to at least one of the epitopes of the protein used to obtain the antibodies. Antibodies of the present invention also include chimeric antibodies that can bind to more than one epitope. Preferred antibodies are raised in response to proteins that are encoded, at least in part, by a flea protease nucleic acid molecule of the present invention.

Anti-flea antibodies of the present invention include antibodies raised in an animal administered a flea protease vaccine of the present invention that exert their effect when fleas feed from the vaccinated animal's blood containing such antibodies. Anti-flea antibodies of the present invention also include antibodies raised in an animal against one or more flea protease proteins, or soluble flea midgut preparations, of the present invention that are then recovered from the animal using techniques known to those skilled in the art. Yet additional antibodies of the present invention are produced recombinantly using techniques as heretofore disclosed for flea protease proteins of the present invention. Antibodies produced against defined proteins can be advantageous because such antibodies are not substantially contaminated with antibodies against other substances that might otherwise cause interference in a diagnostic assay or side effects if used in a therapeutic composition.

Anti-flea protease antibodies of the present invention have a variety of uses that are within the scope of the present invention. For example, such antibodies can be used in a composition of the present invention to passively immunize an animal in order to protect the animal from flea infestation. Anti-flea antibodies can also be used as tools to screen expression libraries and/or to recover desired proteins of the present invention from a mixture of proteins and other contaminants. Furthermore, antibodies of the present invention can be used to target cytotoxic agents to fleas in order to kill fleas. Targeting can be accomplished by conjugating (i.e., stably joining) such antibodies to the cytotoxic agents using techniques known to those skilled in the art.

A preferred anti-flea protease antibody of the present invention can selectively bind to, and preferentially reduce the proteolytic activity of, a flea serine protease, a flea metalloprotease, a flea aspartic acid protease and/or a flea cysteine protease. More preferred anti-flea protease antibodies include anti-flea serine protease antibodies, anti-flea metalloprotease antibodies, anti-flea aminopeptidase antibodies, and anti-flea cysteine protease antibodies. Particularly preferred are anti-flea serine protease antibodies, anti-flea aminopeptidase antibodies, and anti-flea cysteine protease antibodies, including those raised against flea serine protease proteins, flea aminopeptidase proteins or cysteine protease proteins of the present invention.

The present invention also includes the use of protease inhibitors that reduce proteolytic activity of flea proteases to reduce flea infestation of animals and the surrounding environment. As used herein, protease inhibitors are compounds that interact directly with a protease thereby inhibiting that protease's activity, usually by binding to or otherwise interacting with the protease's active site. Protease inhibitors are usually relatively small compounds and as such differ from anti-protease antibodies that interact with the active site of a protease.

Protease inhibitors can be used directly as compounds in compositions of the present invention to treat animals as long as such compounds are not harmful to the animals being treated. Protease inhibitors can also be used to identify preferred types of flea proteases to target using compositions of the present invention. For example, the inventors have shown herein the predominance of serine proteases in flea midguts, particularly in soluble flea midgut preparations, using protease inhibitors. Such knowledge suggests that effective reduction of flea infestation of an animal can be achieved using serine protease vaccines, anti-flea serine protease antibodies and other inhibitors of serine protease synthesis and activity that can be tolerated by the animal. For example, flea immunoglobulin proteinase activity disclosed herein can be targeted to reduce flea infestation. That other proteases are also present in flea midguts according to the present invention also suggests targeting such proteases. Methods to use protease inhibitors are known to those skilled in the art; examples of such methods are disclosed herein.

In one embodiment, a protease inhibitor that can be used in a composition of the present invention to treat an animal is identified by a method including the following steps: (a) identifying candidate (i.e., putative, possible) inhibitor compounds by testing the efficacy of one or more protease inhibitors (i) in vitro for their ability to inhibit flea protease activity and/or (ii) in a flea feeding assay for their ability to reduce the survival and/or fecundity of fleas by adding the inhibitors to the blood meal of a flea being maintained, for example, in a feeding system, such as that described by Wade et al., 1988, J.Med Entomol. 25, 186-190; and (b) testing the efficacy of the candidate inhibitor compounds in animals infested with fleas. Although one does not need both in vitro assay data and flea feeding assay data to determine which candidate compounds to administer to animals, evaluation of both sets of data is preferred since data from neither of the assays necessarily predicts data to be obtained from the other assay. For example, candidate compounds identified using the in vitro assay may work "in the test tube" but may not work in vivo for a number of reasons, including the presence of interfering components in the blood meal that inhibit the activity of such compounds; e.g., although aprotinin can inhibit at least some flea serine proteases in vitro, aprotinin does not work well in the presence of serum proteins, such as are found in the blood. Furthermore, candidate inhibitor compounds identified by the flea feeding assays can include not only desired compounds but also compounds that reduce the viability and/or fecundity of fleas due to general toxicity (e.g., affecting the mitochondria of fleas).

In a preferred embodiment, an inhibitor of a flea protease of the present invention is identified by a method comprising: (a) contacting an isolated flea protease protein comprising an amino acid sequence including SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:16, SEQ ID NO:19, SEQ ID NO:22, SEQ ID NO:24, SEQ ID NO:27, SEQ ID NO:30, SEQ ID NO:33, SEQ ID NO:36, SEQ ID NO:38, SEQ ID NO:41, SEQ ID NO:44, SEQ ID NO:67, SEQ ID NO:68, SEQ ID NO:69, SEQ ID NO:70, SEQ ID NO:71, SEQ ID NO:72, SEQ ID NO:73, SEQ ID NO:96, SEQ ID NO:2, SEQ ID NO:5, SEQ ID NO:8, SEQ ID NO:89, SEQ ID NO:92 and/or SEQ ID NO:9S with a putative inhibitory compound under conditions in which, in the absence of the compound, the protein has proteolytic activity; and (b) determining if the putative inhibitory compound inhibits the activity. A test kit can be used to perform such method. A preferred test kit comprises an isolated flea protease protein comprising an amino acid sequence including SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:16, SEQ ID NO:19, SEQ ID NO:22, SEQ ID NO:24, SEQ ID NO:27, SEQ ID NO:30, SEQ ID NO:33, SEQ ID NO:36, SEQ ID NO:38, SEQ ID NO:41, SEQ ID NO:44, SEQ ID NO:67, SEQ ID NO:68, SEQ ID NO:69, SEQ ID NO:70, SEQ ID NO:71, SEQ ID NO:72, SEQ ID NO:73, SEQ ID NO:96, SEQ ID NO:2, SEQ ID NO:5, SEQ ID NO:8, SEQ ID NO:89, SEQ ID NO:92 and/or SEQ ID NO:95, and a means for determining the extent of inhibition of the activity in the presence of a putative inhibitory compound.

In another embodiment, protease inhibitors are used in the purification of corresponding proteases by, for example, affinity chromatography, in which, a protease inhibitor is incubated with a mixture containing a desired protease under conditions that the inhibitor forms a complex with the protease. The protease can then be recovered from the complex. The protease inhibitor can be attached to a solid support and/or be labelled with, for example, a radioactive, fluorescent, or enzymatic tag that can be used to detect and/or recover the complex.

Suitable protease inhibitors to use in accordance with the present invention include serine protease inhibitors (including immunoglobulin proteinase inhibitors and serpins), metalloprotease inhibitors, aspartic acid protease inhibitors, cysteine protease inhibitors and aminopeptidase inhibitors. Preferred protease inhibitors include serine protease inhibitors, metalloprotease inhibitors, aminopeptidase inhibitors and cysteine protease inhibitors, particularly those that are broad spectrum inhibitors. More preferred are broad spectrum serine protease inhibitors.

There is a wide variety of protease inhibitors, as is known to one skilled in the art. Examples include, but are not limited to, AEBSF, aprotinin, bestatin, chloromethyl ketones TLCK (Nα-p-tosyl-L-lysine chloromethyl ketone) and TPCK (N-tosyl-L-phenylalanine chloromethyl ketone), chymostatin, cystatin, 3'4-dichloroisocoumarin, E-64 (trans-epoxysuccinyl-L-leucylamido-(4-guanidino)butane), EDTA (ethylenediaminetetraacetic acid), leupeptin, methyl ketones having a variety of leaving groups, oxidized L-leucinethiol, pepstatin, 1,10-orthophenanthroline, phosphoramidon, soybean trypsin/chymotrypsin inhibitor and soybean trypsin inhibitor. Preferred protease inhibitors for use in the present invention include AEBSF, bestatin, E-64 leupeptin, pepstatin, 1,10-orthophenanthroline, phosphoramidon, TLCK and TPCK, with AEBSF (a broad spectrum serine protease inhibitor), bestatin (an inhibitor of leucine aminopeptidase) and 1,10-orthophenanthroline (a broad spectrum metalloprotease inhibitor) being particularly preferred.

Protease inhibitors can be produced using methods known to those skilled in the art. Protein- or peptide-based protease inhibitors, such as cystatin or small peptides comprising a protease substrate, can be produced recombinantly and modified as necessary.

The present invention also includes the use of proteolytically active flea protease proteins of the present invention to identify additional protease inhibitors, and preferably protease inhibitor compounds that can be included in a composition of the present invention to be administered to animals. A method to identify a flea protease inhibitor includes the steps of (a) contacting (e.g., combining, mixing) an isolated flea protease protein with a putative (i.e., candidate) inhibitory compound under conditions in which, in the absence of the compound, the protein has proteolytic activity, and (b) determining if the putative inhibitory compound inhibits the proteolytic activity of the protein. Putative inhibitory compounds to screen include organic molecules, antibodies (including functional equivalents thereof) and substrate analogs. Methods to determine protease activity are known to those skilled in the art, as heretofore disclosed. Particularly preferred for use in identifying inhibitors are flea serine protease proteins, flea aminopeptidase proteins and flea cysteine protease proteins of the present invention.

The present invention also includes inhibitors isolated by such a method, and/or test kit, and their use to inhibit any flea protease that is susceptible to such an inhibitor.

It is to be appreciated that the present invention also includes mimetopes of compounds of the present invention that can be used in accordance with methods as disclosed for compounds of the present invention. As used herein, a mimetope of a proteinaceous compound of the present invention (e.g., a flea protease protein, an anti-flea protease antibody, a proteinaceous inhibitor of protease activity or synthesis) refers to any compound that is able to mimic the activity of that proteinaceous compound, often because the mimetope has a structure that mimics the proteinaceous compound. For example, a mimetope of a flea protease protein is a compound that has an activity similar to that of an isolated flea protease protein of the present invention. Mimetopes can be, but are not limited to: peptides that have been modified to decrease their susceptibility to degradation; anti-idiotypic and/or catalytic antibodies, or fragments thereof; non-proteinaceous immunogenic portions of an isolated protein (e.g., carbohydrate structures); and synthetic or natural organic molecules, including nucleic acids. Such mimetopes can be designed using computer-generated structures of proteins of the present invention. Mimetopes can also be obtained by generating random samples of molecules, such as oligonucleotides, peptides or other organic molecules, and screening such samples by affinity chromatography techniques using the corresponding binding partner.

The present invention includes therapeutic compositions, also referred to herein as compositions, that include a (i.e., at least one) compound of the present invention. Preferred compounds to include in a composition of the present invention include flea protease vaccines, anti-flea protease antibodies and/or protease inhibitors as disclosed herein. Such a therapeutic composition can protect an animal from flea infestation by reducing flea protease activity, thereby reducing flea burden on the animal and in the environment of the animal.

Particularly preferred therapeutic compositions of the present invention include at least one of the following compounds: an isolated flea serine protease protein or a mimetope thereof; an isolated flea serine protease nucleic acid molecule that hybridizes under stringent hybridization conditions with a flea serine protease gene; an isolated antibody that selectively binds to a flea serine protease protein; an inhibitor of flea serine protease activity identified by its ability to inhibit flea serine protease activity; an isolated flea cysteine protease protein or a mimetope thereof; an isolated flea cysteine protease nucleic acid molecule that hybridizes under stringent hybridization conditions with a flea cysteine protease gene; an isolated antibody that selectively binds to a flea cysteine protease protein; and an inhibitor of flea cysteine protease activity identified by its ability to inhibit flea cysteine protease activity.

Another embodiment of the present invention is a therapeutic composition that includes a first compound that reduces flea protease activity and a second compound that reduces flea burden by a method other than by reducing flea protease activity. The present invention also includes a method to protect an animal from flea infestation by administering to the animal such a composition. The first compound of such a composition by effectively reducing flea protease activity in the midgut, enhances the activity of the second compound. While not being bound by theory, it is believed that a number of anti-flea treatments, particularly those that are proteinaceous, are not very effective because they are degraded in the flea midgut. The present invention permits the effective use of such anti-flea treatments by reducing proteolytic degradation of such treatments by the flea midgut.

Preferred first compounds to include in such a composition include flea protease vaccines, anti-flea protease antibodies and/or protease inhibitors as disclosed herein, such compounds that target flea immunoglobulin proteinase activity.

A preferred therapeutic composition of the present invention comprises an excipient and a protective compound including: an isolated protein or mimetope thereof encoded by a nucleic acid molecule that hybridizes under stringent hybridization conditions with a nucleic acid molecule having a nucleic acid sequence encoding a protein comprising an amino acid sequence including SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:16, SEQ ID NO:19, SEQ ID NO:22, SEQ ID NO:24, SEQ ID NO:27, SEQ ID NO:30, SEQ ID NO:33, SEQ ID NO:36, SEQ ID NO:38, SEQ ID NO:41, SEQ ID NO:44, SEQ ID NO:67, SEQ ID NO:68, SEQ ID NO:69, SEQ ID NO:70, SEQ ID NO:71, SEQ ID NO:72, SEQ ID NO:73, SEQ ID NO:96, SEQ ID NO:2, SEQ ID NO:5, SEQ ID NO:8, SEQ ID NO:89, SEQ ID NO:92 and/or SEQ ID NO:95; an isolated nucleic acid molecule that hybridizes under stringent hybridization conditions with a gene comprising a nucleic acid sequence including SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:23, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:28, SEQ ID NO:29, SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:34, SEQ ID NO:35, SEQ ID NO:37, SEQ ID NO:39, SEQ ID NO:40, SEQ ID NO:42, SEQ ID NO:43, SEQ ID NO:45, SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:88, SEQ ID NO:90, SEQ ID NO:91, SEQ ID NO:93 and/or SEQ ID NO:94; an isolated antibody that selectively binds to a protein encoded by a nucleic acid molecule that hybridizes under stringent hybridization conditions with a nucleic acid molecule having a nucleic acid sequence encoding a protein comprising an amino acid sequence including SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:16, SEQ ID NO:19, SEQ ID NO:22, SEQ ID NO:24, SEQ ID NO:27, SEQ ID NO:30, SEQ ID NO:33, SEQ ID NO:36, SEQ ID NO:38, SEQ ID NO:41, SEQ ID NO:44, SEQ ID NO:67, SEQ ID NO:68, SEQ ID NO:69, SEQ ID NO:70, SEQ ID NO:71, SEQ ID NO:72, SEQ ID NO:73, SEQ ID NO:96, SEQ ID NO:2, SEQ ID NO:5, SEQ ID NO:8, SEQ ID NO:89, SEQ ID NO:92 and/or SEQ ID NO:95; an inhibitor of protease activity identified by its ability to inhibit the activity of a protein encoded by a nucleic acid molecule that hybridizes under stringent hybridization conditions with a nucleic acid molecule having a nucleic acid sequence encoding a protein comprising an amino acid sequence including SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:16, SEQ ID NO:19, SEQ ID NO:22, SEQ ID NO:24, SEQ ID NO:27, SEQ ID NO:30, SEQ ID NO:33, SEQ ID NO:36, SEQ ID NO:38, SEQ ID NO:41, SEQ ID NO:44, SEQ ID NO:67, SEQ ID NO:68, SEQ ID NO:69, SEQ ID NO:70, SEQ ID NO:71, SEQ ID NO:72, SEQ ID NO:73, SEQ ID NO:96, SEQ ID NO:2, SEQ ID NO:5, SEQ ID NO:8, SEQ ID NO:89, SEQ ID NO:92 and/or SEQ ID NO:95; and a mixture thereof.

Suitable second compounds include any anti-flea agent(s), including, but not limited to, proteinaceous compounds, insecticides and flea collars. Preferred second compounds are proteinaceous compounds that effect active immunization (e.g., antigen vaccines), passive immunization (e.g., antibodies), or that otherwise inhibit a flea activity that when inhibited can reduce flea burden on and around an animal. Examples of second compounds include a compound that inhibits binding between a flea membrane protein and its ligand (e.g., a compound that inhibits flea ATPase activity or a compound that inhibits binding of a peptide or steroid hormone to its receptor), a compound that inhibits hormone (including peptide or steroid hormones) synthesis, a compound that inhibits vitellogenesis (including production of vitellin and transport and maturation thereof into a major egg yolk protein), a compound that inhibits fat body function, a compound that inhibits flea muscle action, a compound that inhibits the flea nervous system, a compound that inhibits the flea immune system and/or a compound that inhibits flea feeding.

Compositions of the present invention can also include other components such as a pharmaceutically acceptable excipient, an adjuvant, and/or a carrier. For example, compositions of the present invention can be formulated in an excipient that the animal to be treated can tolerate.

Examples of such excipients include water, saline, Ringer's solution, dextrose solution, Hank's solution, and other aqueous physiologically balanced salt solutions. Nonaqueous vehicles, such as fixed oils, sesame oil, ethyl oleate, or triglycerides may also be used. Other useful formulations include suspensions containing viscosity enhancing agents, such as sodium carboxymethylcellulose, sorbitol, or dextran. Excipients can also contain minor amounts of additives, such as substances that enhance isotonicity and chemical stability. Examples of buffers include phosphate buffer, bicarbonate buffer and Tris buffer, while examples of preservatives include thimerosal, m- or o-cresol, formalin and benzyl alcohol. Standard formulations can either be liquid injectables or solids which can be taken up in a suitable liquid as a suspension or solution for injection. Thus, in a non-liquid formulation, the excipient can comprise dextrose, human serum albumin, preservatives, etc., to which sterile water or saline can be added prior to administration.

In one embodiment of the present invention, the composition can also include an immunopotentiator, such as an adjuvant or a carrier. Adjuvants are typically substances that generally enhance the immune response of an animal to a specific antigen. Suitable adjuvants include, but are not limited to, Freund's adjuvant; other bacterial cell wall components; aluminum-based salts; calcium-based salts; silica; polynucleotides; toxoids; serum proteins; viral coat proteins; other bacterial-derived preparations; gamma interferon; block copolymer adjuvants, such as Hunter's Titermax adjuvant (Vaxcel™, Inc. Norcross, Ga.); Ribi adjuvants (available from Ribi ImmunoChem Research, Inc., Hamilton, Mont.); and saponins and their derivatives, such as Quil A (available from Superfos Biosector A/S, Denmark). Carriers are typically compounds that increase the half-life of a therapeutic composition in the treated animal. Suitable carriers include, but are not limited to, polymeric controlled release formulations, biodegradable implants, liposomes, bacteria, viruses, oils, esters, and glycols.

One embodiment of the present invention is a controlled release formulation that is capable of slowly releasing a composition of the present invention into an animal. As used herein a controlled release formulation comprises a composition of the present invention in a controlled release vehicle. Suitable controlled release vehicles include, but are not limited to, biocompatible polymers, other polymeric matrices, capsules, microcapsules, microparticles, bolus preparations, osmotic pumps, diffusion devices, liposomes, lipospheres, and transdermal delivery systems. Other controlled release formulations of the present invention include liquids that, upon administration to an animal, form a solid or a gel in situ. Preferred controlled release formulations are biodegradable (i.e., bioerodible).

A preferred controlled release formulation of the present invention is capable of releasing a composition of the present invention into the blood of the treated animal at a constant rate sufficient to attain therapeutic dose levels of the composition to reduce protease activity in fleas feeding from the animal over a period of time ranging from about 1 to about 12 months. A controlled release formulation of the present invention is capable of effecting a treatment for preferably at least about 1 month, more preferably at least about 3 months and even more preferably for at least about 6 months, even more preferably for at least about 9 months, and even more preferably for at least about 12 months.

In order to protect an animal from flea infestation, a therapeutic composition of the present invention is administered to the animal in an effective manner such that the protease activity of fleas feeding from the blood stream of animals treated with the composition is reduced. As such, a treated animal is an animal that is competent to reduce the flea burden by reducing flea protease activity, or by reducing flea protease activity and at least one other flea activity. Preferably, the protease activity is reduced by at least about 50 percent, more preferably by at least about 70 percent and even more preferably by at least about 90 percent. Methods to administer compositions to the animal in order to render the animal competent depend on the nature of the composition and administration regime. Animals administered a protease vaccine with at least one booster shot usually become competent at about the same time as would be expected for any vaccine treatment. For example, animals administered a booster dose about 4 to 6 weeks after a primary dose usually become competent within another about 3 to 4 weeks. Animals administered a composition including an anti-flea protease antibody or protease inhibitor become competent as soon as appropriate serum levels of the compound are achieved, usually with one to three days.

In a preferred embodiment, a composition of the present invention when administered to a host animal is able to reduce flea viability by at least about 50 percent within at least about 21 days after the fleas begin feeding from the treated animal. (Note that fleas usually live about 40 days to about 50 days on one or more animals.) A more preferred composition when administered to a host animal is able to reduce flea viability by at least about 65 percent within at least about 14 days after the fleas begin feeding from the treated animal. An even more preferred composition when administered to an animal is able to reduce flea viability by at least about 90 percent within at least about 7 days after the fleas begin feeding from the treated animal.

In another preferred embodiment, a composition of the present invention when administered to a host animal is able to reduce flea fecundity (i.e., egg laying ability) by at least about 50 percent, more preferably by at least about 70 percent, and even more preferably by at least about 90 percent, within at least about 30 days after the fleas begin feeding from the treated animal. (Note that fleas usually do not begin laying eggs until about 7 days after taking a blood meal.)

In accordance with the present invention, compositions are administered to an animal in a manner such that the animal becomes competent to reduce flea protease activity in a flea that feeds from the competent; i.e., the animal becomes a treated animal. For example, a flea protease vaccine of the present invention, when administered to an animal in an effective manner, is able to elicit (i.e., stimulate) an immune response that produces an antibody titer in the blood stream of the animal sufficient to reduce flea protease activity. Similarly, an anti-flea protease antibody of the present invention, when administered to an animal in an effective manner, is administered in an amount so as to be present in the animal's blood stream at a titer that is sufficient to reduce flea protease activity. A protease inhibitor compound of the present invention, when administered to an animal in an effective manner, is administered in a manner so as to be present in the animal's blood stream at a concentration that is sufficient to reduce flea protease activity. Oligonucleotide nucleic acid molecules of the present invention can also be administered in an effective manner, thereby reducing expression of flea proteases.

Compositions of the present invention can be administered to animals prior to or during flea infestation. It is to be noted that when vaccines of the present invention are administered to an animal, a time period is required for the animal to elicit an immune response before the animal is competent to inhibit protease activity of fleas feeding from that animal. Methods to obtain an immune response in an animal are known to those skilled in the art.

Acceptable protocols to administer compositions in an effective manner include individual dose size, number of doses, frequency of dose administration, and mode of administration. Determination of such protocols can be accomplished by those skilled in the art. A suitable single dose is a dose that is capable of protecting an animal from flea infestation when administered one or more times over a suitable time period. For example, a preferred single dose of a protease vaccine or a mimetope thereof ranges from about 1 microgram (μg, also denoted ug) to about 10 milligrams (mg) of the composition per kilogram body weight of the animal. Booster vaccinations can be administered from about 2 weeks to several years after the original administration. Booster vaccinations preferably are administered when the immune response of the animal becomes insufficient to protect the animal from flea infestation. A preferred administration schedule is one in which from about 10 μg to about 1 mg of the vaccine per kg body weight of the animal is administered from about one to about two times over a time period of from about 2 weeks to about 12 months. In one embodiment, a booster dose of a composition of the present invention is administered about 4 to 6 weeks after the primary dose, and additional boosters are administered about once or twice a year. Modes of administration can include, but are not limited to, oral, nasal, topical, transdermal, rectal, and parenteral routes. Parenteral routes can include, but are not limited to subcutaneous, intradermal, intravenous, and intramuscular routes.

In another embodiment, a preferred single dose of an anti-flea protease antibody composition or a mimetope thereof ranges from about 1 μg to about 10 mg of the composition per kilogram body weight of the animal. Anti-flea antibodies can be re-administered from about 1 hour to about biweekly for several weeks following the original administration. Booster treatments preferably are administered when the titer of antibodies of the animal becomes insufficient to protect the animal from flea infestation. A preferred administration schedule is one in which from about 10 μg to about 1 mg of an anti-flea protease antibody composition per kg body weight of the animal is administered about every 2 to every 4 weeks. Suitable modes of administration are as disclosed herein and are known to those skilled in the art.

According to one embodiment, a nucleic acid molecule of the present invention can be administered to an animal in a fashion to enable expression of that nucleic acid molecule into a protective protein (e.g., flea protease vaccine, anti-flea protease antibody, or proteinaceous protease inhibitor) or protective RNA (e.g., antisense RNA, ribozyme or RNA drug) in the animal to be protected from disease. Nucleic acid molecules can be delivered to an animal in a variety of methods including, but not limited to, (a) direct injection (e.g., as "naked" DNA or RNA molecules, such as is taught, for example in Wolff et al., 1990, Science 247, 1465-1468) or (b) packaged as a recombinant virus particle vaccine or as a recombinant cell vaccine (i.e., delivered to a cell by a vehicle selected from the group consisting of a recombinant virus particle vaccine and a recombinant cell vaccine).

A recombinant virus particle vaccine of the present invention includes a recombinant molecule of the present invention that is packaged in a viral coat and that can be expressed in an animal after administration. Preferably, the recombinant molecule is packaging-deficient. A number of recombinant virus particles can be used, including, but not limited to, those based on alphaviruses, poxviruses, adenoviruses, herpesviruses, and retroviruses. When administered to an animal, a recombinant virus particle vaccine of the present invention infects cells within the immunized animal and directs the production of a protective protein or RNA nucleic acid molecule that is capable of protecting the animal from disease caused by a parasite of the present invention. A preferred single dose of a recombinant virus particle vaccine of the present invention is from about 1×10⁴ to about 1×10⁷ virus plaque forming units (pfu) per kilogram body weight of the animal. Administration protocols are similar to those described herein for protein-based vaccines.

A recombinant cell vaccine of the present invention includes recombinant cells of the present invention that express at least one protein of the present invention. Preferred recombinant cells include Salmonella, E. coli, Mycobacterium, S. frugiperda, baby hamster kidney, myoblast G8, COS, MDCK and CRFK recombinant cells, with Salmonella recombinant cells being more preferred. Such recombinant cells can be administered in a variety of ways but have the advantage that they can be administered orally, preferably at doses ranging from about 10⁸ to about 10¹² bacteria per kilogram body weight. Administration protocols are similar to those described herein for protein-based vaccines. Recombinant cell vaccines can comprise whole cells or cell lysates.

Compositions of the present invention can be administered to any animal susceptible to flea infestation, including warm-blooded animals. Preferred animals to treat include mammals and birds, with cats, dogs, humans, cattle, chinchillas, ferrets, goats, mice, minks, rabbits, raccoons, rats, sheep, squirrels, swine, chickens, ostriches, quail and turkeys as well as other furry animals, pets and/or economic food animals, being more preferred. Particularly preferred animals to protect are cats and dogs.

The present invention includes compositions to treat flea infestation by any flea. As such, compositions of the present invention can be derived from any flea species. Preferred fleas to target include fleas of the following genera: Ctenocephalides, Cyopsyllus, Diamanus (Oropsylla), Echidnophaga, Nosopsyllus, Pulex, Tunga, and Xenopsylla, with those of the species Ctenocephalides canis, Ctenocephalides felis, Diamanus montanus, Echidnophaga gallinacea, Nosopsyllus faciatus, Pulex irritans, Pulex simulans, Tunga penetrans and Xenopsylla cheopis being more preferred. Particularly preferred fleas from which to protect animals include fleas of the species Ctenocephalides felis, Ctenocephalides canis, and Pulex species (e.g., Pulex irritans and Pulex simulans). It is also within the scope of the present invention to administer compositions of the present invention directly to fleas.

The present invention also includes the use of compositions of the present invention to reduce infestation by other ectoparasites as well as the use of compositions including protease vaccines, anti-protease antibodies and compounds that inhibit protease synthesis and/or activity derived from any ectoparasite to reduce ectoparasite infestation, particularly controlled release formulations containing such compositions. Preferred ectoparasites to target include arachnids, insects and leeches. More preferred ectoparasites to target include fleas; ticks, including both hard ticks of the family Ixodidae (e.g., Ixodes and Amblyomma) and soft ticks of the family Argasidae (e.g., Ornithodoros, such as O. parkeri and O. turicata); flies, such as midges (e.g., Culicoides), mosquitos, sand flies, black flies, horse flies, horn flies, deer flies, tsetse flies, stable flies, myiasis-causing flies and biting gnats; ants; spiders, lice; mites; and true bugs, such as bed bugs and kissing bugs, including those carrying Chagas disease. Even more preferred ectoparasites to target include fleas, mosquitos, midges, sandflies, blackflies, ticks and Rhodnius.

The following examples are provided for the purposes of illustration and are not intended to limit the scope of the present invention.

EXAMPLES

It is to be noted that the Examples include a number of molecular biology, microbiology, immunology and biochemistry techniques considered to be known to those skilled in the art. Disclosure of such techniques can be found, for example, in Sambrook et al., ibid., Borovsky, Arch Insect Biochem. and Phys., 7:187-210, 1988, and related references. Examples 1 through 21, and the the sequence information provided in the sequence listing therein, of related PCT Publication No. WO 96/11706, published Apr. 25, 1996, are incorporated herein by this reference in their entirety.

Example 1

This example describes the cloning and sequencing of a flea cysteine protease nucleic acid molecule.

A flea cysteine protease nucleic acid molecule, referred to herein as nfCP1₅₇₃ was produced by PCR amplification using the following method. Primer Cal3F (designed to obtain a calreticulin gene), having nucleic acid sequence 5' TTG GGA TAC ACT TTG ACT GTT AAC C 3', represented herein as SEQ ID NO:97 was used in combination with the M13 universal primer, to PCR amplify, using standard techniques, a DNA fragment from a bovine blood-fed whole flea cDNA expression library as described above in Example 8 of related PCT Publication No. WO 96/11706. Surprisingly, the isolated DNA fragment correlated with a cysteine protease nucleic acid sequence. Sequence from this DNA fragment was used to design primer Cys1R, having the nucleic acid sequence 5' GTG AGC AAC CAT TAT TTC CAT ATC 3', represented herein as SEQ ID NO:98, which was used in a second PCR amplification in combination with the M13 reverse primer. A third PCR amplification was performed using primer Cys1F, having the nucleic acid sequence 5' CTT TCC TCA CAA TAC CAC CAA GGA AGC 3', represented herein as SEQ ID NO:74, in combination with the M13 universal primer. A fourth PCR amplification was performed using primer Cys2F, having the nucleic acid sequence 51 CTT GTA CGA TTG TCT CAA CAG GC 3', represented herein as SEQ ID NO:76, in combination with the M13 universal primer. The resulting PCR products were each gel purified and cloned into the TA Vector® System, and subjected to standard DNA sequencing techniques. A composite nucleic acid sequence representing a flea cysteine protease coding region was deduced, referred to herein as nfCP1₅₇₃, was deduced and is denoted herein as SEQ ID NO:76. Translation of SEQ ID NO:76 suggests that nucleic acid molecule nfCP1₅₇₃ encodes a non-full-length flea cysteine protease protein of about 191 amino acids, referred to herein as PfCP1₁₉₁, having amino acid sequence SEQ ID NO:77, assuming the first codon spans from about nucleotide 1 through about nucleotide 3 of SEQ ID NO:76.

The nucleic acid and amino acid sequences of the nfCP1₅₇₃ nucleic acid molecule and PfCP1₁₉₁ protein, respectively, were compared to known nucleic acid and amino acid sequences using a Genbank homology search. SEQ ID NO:77 was found to be similar to the amino acid sequence of P. sativum cysteine protease. The most highly conserved region of continuous similarity between SEQ ID NO:77 and P. sativum cysteine protease amino acid sequences spans from about amino acid 71 through about amino acid 165 of SEQ ID NO:77 and from about amino acid 17 through about amino acid 168 of the P. sativum cysteine protease, there being about 42% identity between the two regions. Comparison of the nucleic acid sequence encoding amino acids from about 205 through about 492 of nfCP1₅₇₃ indicate that those regions are about 54% identical.

Example 2

This example describes the cloning and sequencing of certain flea serine protease nucleic acid molecules.

Certain serine protease cDNA nucleic acid molecules have been isolated from reverse transcriptase PCR amplification of mRNA isolated from cat blood-fed whole fleas. The mRNA was isolated from fleas gathered over 72 hours after the initiation of feeding on cat blood. As such, the mRNA comprised a mixture of mRNA isolated at different time points over 72 hours. The mRNA was isolated using ground-up fleas, extracting total flea RNA using Tri-Reagent (available from Molecular Research Center, Cincinnati, Ohio) and an Invitrogen Fast Track™ RNA isolation kit (available from Invitrogen, Inc. San Diego, Calif.). cDNA was synthesized using a Stratagene RT-PCR kit (available from Stratagene, Inc, San Diego, Calif.). Primers used for first-strand cDNA synthesis included an equal molar mixture of the following: 5'dT-2VT3' and 5'dT-2VC3' (as provided in a differential display kit, available from Operon Technologies, Inc. Alameda, Calif.).

The actual primers used in the PCR amplification of the cDNA described above included cat-try #2 (SEQ ID NO:86) used in combination with H57 primer (SEQ ID NO:99). The resultant PCR products were gel purified and cloned into the TA Vector™. Recombinant TA vector clones were isolated and the nucleic acid molecules were subjected to nucleic acid sequencing using analysis as described above.

A. A nucleic acid sequence of a flea serine protease nucleic molecule, namely nfSP24₄₁₀, is represented herein as SEQ ID NO:78. Translation of SEQ ID NO:78. suggests that nucleic acid molecule nfSP24₄₁₀ encodes a non-full-length flea serine protease protein of about 136 amino acids, referred to herein as PfSP24₁₃₆, having amino acid sequence SEQ ID NO:79, assuming the first codon spans from about nucleotide 1 through about nucleotide 3 of SEQ ID NO:78. A Genbank homology search revealed most homology between SEQ ID NO:79 and an Anopheles gambiae chymotrypsin protein sequence, there being about 38% identity between corresponding regions of the two amino acid sequences.

B. Another nucleic acid sequence of a flea serine protease nucleic molecule, namely nfSP33₄₂₆, is represented herein as SEQ ID NO:82. Translation of SEQ ID NO:82 suggests that nucleic acid molecule nfSP33₄₂₆ encodes a non-full-length flea serine protease protein of about 142 amino acids, referred to herein as PfSP33₁₄₂, having amino acid sequence SEQ ID NO:83, assuming the first codon spans from about nucleotide 1 through about nucleotide 3 of SEQ ID NO:82. A Genbank homology search revealed most homology between SEQ ID NO:83 and a Drosophila serine protease stubble protein sequence, there being about 45% identity between corresponding regions of the two amino acid sequences.

Example 3

This example describes the cloning and sequencing of a flea serine protease nucleic acid molecule.

A serine protease cDNA nucleic acid molecule was isolated in a manner similar to that described in Example 8 of related PCT Publication No. WO 96/11706. The actual primers used in PCR amplification of the serine protease nucleic acid molecule from a cat blood-fed whole flea cDNA expression library (produced as described in Example 2) included cat-try #2 (SEQ ID NO:86) in combination with M13 reverse primer (SEQ ID NO:87). The resulting PCR product was diluted 1:25 and used as a template in a second PCR reaction using the forward vector primer T3 in combination with the reverse primer (derived from the nucleic acid sequence of nfSP33₇₇₈, described in Example 2) having the nucleic acid sequence 5' ATT CCT CGT GGT TCA GTC GCT C 3', represented herein as SEQ ID NO:100. The resultant PCR product was gel purified and cloned into the TA Vector™. The clones were subjected to nucleic acid sequencing as described above.

A nucleic acid sequence of a flea serine protease nucleic molecule, namely nfSP33₇₇₈ is represented herein as SEQ ID NO:84. As expected, SEQ ID NO:84 includes a portion of SEQ ID NO:82 Translation of SEQ ID NO:84 suggests that nucleic acid molecule nfSP33₇₇₈ encodes a non-full-length flea serine protease protein of about 259 amino acids, referred to herein as PfSP33₂₅₉, having amino acid sequence SEQ ID NO:85, assuming the first codon spans from about nucleotide 2 through about nucleotide 4 of SEQ ID NO: 84. A Genbank homology search revealed most homology between SEQ ID NO: 84 and a Drosophila serine protease stubble gene, there being about 54% identity between nucleotides 23-778 of SEQ ID NO:84 and nucleotides 2324-3064 of the Drosophila serine protease stubble gene.

Example 4

This example describes the cloning and sequencing of certain larval flea serine protease nucleic acid molecules.

Certain serine protease cDNA nucleic acid molecules have been isolated from a mixed instar larval cDNA library produced using 1st, 2nd and 3rd instar larvae fed on cat blood, by PCR amplification. The actual primers used in the PCR amplification included either cat-try #2 (SEQ ID NO:86) in combination with either H57 primer (SEQ ID NO:99) or M13 reverse primer (SEQ ID NO:87). The resultant PCR products were gel purified and cloned into the TA Vector™. Three recombinant TA vector clones were isolated containing PCR products using cat-try #2 and M13 reverse as primers and one clone was isolated containing PCR products using cat-try #2 and H57 primers. These newly cloned nucleic acid molecules were subjected to nucleic acid sequencing as described above. A. A nucleic acid sequence of a larval flea serine protease nucleic molecule isolated using cat-try #2 and H57 primers, namely nfSP32₄₃₃, is represented herein as SEQ ID NO:80. Translation of SEQ ID NO:80 suggests that nucleic acid molecule nfSP32₄₃₃ encodes a non-full-length flea serine protease protein of about 144 amino acids, referred to herein as PfSP32₁₄₄, having amino acid sequence SEQ ID NO:81, assuming the first codon spans from about nucleotide 1 through about nucleotide 3 of SEQ ID NO:80. A Genbank homology search revealed most homology between SEQ ID NO:80 and an Anopheles gambiae trypsin gene, there being about 52% identity between corresponding regions of the two nucleic acid molecules.

Example 5

This example describes the isolation and characterization of a 31 kD flea serine protease.

Guts from about 1500 fleas that had been fed on cat blood for about 24 hours were dissected in Gut Dissection Buffer (50 mM Tris 8.0, 100 mM CaCl₂). The guts were disrupted by freezing and thawing 4 times, followed by sonication. The resulting extracts were clarified by centrifugation for 20 minutes at 14,000 rpm in a microfuge at 4° C. The supernatant was recovered.

The gut supernatant was loaded onto a 3 ml column comprising p-aminobenzamidine cross-linked to Sepharose beads (Sigma), previously equilibrated in Benzamidine Column Buffer (50 mM Tris 8.0, 100 mM CaCl₂, 400 mM NaCl). The supernatant was incubated on the column for about 10 min. Unbound protein was slowly washed off the column using Benzamidine Column Buffer until no protein was detectable by Bradford Assay (Bio Rad).

Proteases bound to the benzamidine column were then eluted using 10 ml Benzamidine Column Buffer supplemented with 10 mM p-aminobenzamidine (brought to pH 8.0 with NaOH). Proteases in the eluant were concentrated and diafiltered into a volume of about 0.3 ml Gut Dissection Buffer using a Microcon 3 concentrator (Amicon).

About 120 μl of the concentrated eluant was further concentrated to a volume of about 30 μl. Proteases contained in this concentrate were resolved by gel electrophoresis on a 14% Tris-Glycine electrophoresis gel (15 μl per lane=approximately 300 gut equivalents per lane). After electrophoresis, the separated proteases were blotted onto a PVDF membrane using a CAPS buffer (10 mM CAPS pH 11, 0.5 mM DTT) The membrane was stained with Coomassie Brilliant Blue. A dominant protein band of about 31 kDa was visualized. The membrane was then used for automated N-terminal sequencing (described in Example 7 of related PCT Publication No. WO 96/11706). A partial N-terminal amino acid sequence of the flea protease was determined to be IVGGEDVDISTCGWC (denoted SEQ ID NO:68).

Example 6

This example describes the isolation and characterization of a 31 kD flea serine protease contained in a formulation having IgGase activity (i.e., ability to proteolyze immunoglobulin G proteins).

Cat blood-fed flea gut extracts were prepared and selected on a benzamidine column as described above in Example 5. IgG protease activity was assayed by incubating at 37° C., overnight, the benzamidine eluant with cat immunoglobulin G proteins (IgG) purified on Protein A sepharose. The ability of the flea gut benzamidine eluant to digest cat IgG was detected by resolving the samples by gel electrophoresis through a 14% SDS-PAGE gel and silver staining the gel using standard methods. The marked decrease (compared with control samples lacking protease activity) of a 50 kDa band on the silver stained gel, representing cat IgG heavy chain, indicated that the benzamidine eluant contains IgG protease activity.

The benzamidine eluant was then purified on a PolyPropylaspartamide hydrophobic interaction chromatography (HIC) column by applying the eluant to the column in buffer containing 0.1 M KPO₄, pH 6.5 and 2 M (N₄ H₂) S₄ O. Proteases bound to the column were eluted using an ammonium sulfate gradient of 2 M to 0 M in HIC column buffer. Column fractions were tested for IgG protease activity using the method described above. Fractions containing IgG protease activity were pooled and applied to a PolyCat cation exchange column in 20 M sodium acetate, pH 6. The proteins were eluted using a sodium chloride gradient of 0 M to 1 M NaCl in 20 M sodium acetate. Fractions eluted from the column were tested for IgG protease activity and then each fraction was resolved by electrophoresis using SDS-PAGE. Fractions having the highest levels of IgG protease activity included a protein band that migrated at about 31 kDa on the SDS-PAGE gel. Weaker protease activity corresponded to an about 28 kDa band.

The 31 kDa protein present on the SDS-PAGE gel was used for N-terminal amino acid sequencing using the blotting method described above. A partial N-terminal amino acid sequence was determined to be IVGGEDVDIST(C)GWQI(S)FQ(S)ENLHF(C)GG(S)IIAPK (denoted herein as SEQ ID NO:69). A comparison of SEQ ID NO:69 and SEQ ID NO:68 (described in Example 5) indicates a single residue difference between the two amino acid sequences at residue 15 of each sequence (i.e., Q and V, respectively). Since SEQ ID NO:69 correlates with IgGase activity, the data suggests that the larval protein containing SEQ ID NO:68 has IgGase activity.

Example 7

This example describes the cloning and sequencing of a 31 kDa flea serine protease contained in a formulation having IgGase activity.

A flea protease nucleic acid molecule was isolated from a cat blood-fed whole flea library (described in Example 2) and a bovine blood-fed whole flea library (described in Example 8 of related PCT Publication No. WO 96/11706) by PCR amplification. The actual primers used in the PCR amplification included FP31A primer designed using the N-terminal amino acid sequence SEQ ID NO:68, the primer having the nucleic acid sequence 5' GAA GAT GTW GAT ATT TCW ACA TGT GG 3' (SEQ ID NO:101) used in combination with the M13 universal primer. The resultant PCR products were gel purified and cloned into the TA Vector™ and subjected to nucleic acid sequencing as described above.

A FP31B primer (5' GAA AAT GAA ATC CAC TTA AAC ATT ACG 3'), (represented herein as SEQ ID NO:102) was designed using the DNA sequence of a DNA fragment from a bovine blood-fed cDNA library. A flea protease cDNA nucleic acid molecule was isolated by PCR amplification of the cat blood-fed whole flea library and the bovine blood-fed whole flea library described above by PCR amplification. PCR amplification was performed using the FP31B primer in combination with M13 reverse primer. The resulting PCR products were then diluted 1:25, and used as a template for a second PCR reaction using primer FP31C, having the sequence 5' CTC TTA TTG TAC GAG GGA TGC 3' (denoted herein SEQ ID NO:103) in combination with T3 primer. The resulting nested PCR product was cloned into TA Vectors and subjected to DNA sequencing.

The nucleic acid sequence of the resulting flea serine protease nucleic molecule, namely nfSP28₉₂₃ is represented herein as SEQ ID NO:66. Translation of SEQ ID NO:66 yields a protein of about 267 amino acids, denoted PfSP28₂₆₇, having amino acid sequence SEQ ID NO:67, assuming an open reading frame in which the putative start codon spans from about nucleotide 8 through about nucleotide 10 of SEQ ID NO:66 or from about nucleotide 11 through about nucleotide 13, and a stop codon spanning from about nucleotide 803 through about nucleotide 805 of SEQ ID NO:66. SEQ ID NO:67 contains SEQ ID NO:68 except Q is substituted for C, and SEQ ID NO:69. A Genbank homology search revealed most homology between SEQ ID NO:66 and Bombix mori vitellin-degrading protease gene, there being about 53% identity between corresponding regions of the two nucleic acid sequences.

Example 8

This example provides additional nucleic acid and deduced amino acid sequences of nucleic acid molecules encoding a flea cysteine protease protein of the present which was described in Example 1. This example also provides the production of a cysteine protease protein in E. coli cells.

A. Additional Cysteine Protease Nucleic Acid Molecule

The PCR products described in Example 1 were submitted to additional nucleic acid sequence analysis in order to obtain the nucleic acid sequence of additional portions of the coding region of the cysteine protease gene. A composite nucleic acid sequence representing a flea cysteine protease coding region, referred to herein as nfCP1₁₁₀₉, was deduced and is denoted herein as SEQ ID NO:1. SEQ ID NO:76 is contained within the sequence of the nucleic acid molecule nfCP1₁₁₀₉. Translation of SEQ ID NO:1 suggests that nucleic acid molecule nfCP1₁₁₀₉ encodes a full-length flea cysteine protease protein of about 327 amino acids, referred to herein as PfCP1₃₂₇, having amino acid sequence SEQ ID NO:2, assuming an open reading frame in which the initiation codon spans from about nucleotide 126 through about nucleotide 128 of SEQ ID NO:1 and the termination codon spans from about nucleotide 1107 through about nucleotide 1109 of SEQ ID NO:1. The complement of SEQ ID NO:1 is represented herein by SEQ ID NO:3. The coding region encoding PfCP1₃₂₇₁ is represented by nucleic acid molecule nfCP1₉₈₄, having a coding strand with the nucleic acid sequence represented by SEQ ID NO:4 and a complementary strand with nucleic acid sequence SEQ ID NO:6. The proposed mature protein, denoted herein as PfCP1₂₂₆, contains about 226 amino acids which is represented herein as SEQ ID NO:8. The nucleic acid molecule encoding PfCP1₂₂₆ is denoted herein as nfCP1₆₈₁, which is represented by SEQ ID NO:7. The amino acid sequence of PfCP1₃₂₇ (i.e., SEQ ID NO:2) predicts that PfCP1₃₂₇ has an estimated molecular weight of about 42 kD and an estimated pI of about pI 8.84.

Comparison of nucleic acid sequence SEQ ID NO:1 with nucleic acid sequences reported in GenBank indicates that SEQ ID NO:1 showed the most homology, i.e., about 55% identity, with the following three genes: a Drosophila cysteine protease gene, a Bombyx cysteine protease gene and a Sarcophaga cysteine protease gene. Comparison of amino acid sequence SEQ ID NO:2 (i.e., the amino acid sequence of PfCP1₃₂₇) with amino acid sequences reported in GenBank indicates that SEQ ID NO:2 showed the most homology, i.e., about 42% identity, with the following three proteins: a Drosophila cysteine protease protein, a Bombyx cysteine protease protein and a Sarcophaga cysteine protease protein.

B. Production of Cysteine Protease Protein in E. coli Cells

An about 660-bp nucleic acid molecule, referred to herein as nfCP1₆₆₀ (designed to encode an apparently mature cysteine protease protein) was PCR-amplified from a flea mixed instar cDNA library produced using unfed 1st instar, bovine blood-fed 1st instar, bovine blood-fed 2^(nd) instar and bovine blood-fed 3^(rd) instar flea larvae (this combination of tissues is referred to herein as mixed instar larval tissues for purposes of this example). Total RNA was extracted from mixed instar tissue using an acid-guanidinium-phenol-chloroform method similar to that described by Chomczynski et al., 1987, Anal. Biochem. 162, p. 156-159. Approximately 5,164 mixed instar larvae were used in each RNA preparation. Poly A+ selected RNA was separated from each total RNA preparation by oligo-dT cellulose chromatography using Poly(A)Quick® mRNA isolation kits (available from Stratagene Cloning Systems, La Jolla, Calif.), according to the method recommended by the manufacturer. A mixed instar cDNA expression library was constructed in lambda (λ) Uni-ZAP™XR vector (available from Stratagene Cloning Systems) using Stratagene's ZAP-cDNA Synthesis Kit® protocol. About 6.34 μg of mixed instar poly A+ RNA were used to produce the mixed instar library. The resultant mixed instar library was amplified to a titer of about 2.17×10¹⁰ pfu/ml with about 97% recombinants. The primers used in the PCR amplification were sense primer CysBS' having the nucleotide sequence 5' GAT AAG GAT CCG TTA CCA GAT TCT TTC GAC TGG 3' (containing a BamHI-site; denoted SEQ ID NO:64) and anti-sense primer CysHA having the nucleotide sequence 5' TTA TCA AGC TTC CAT TTA CAT GCC GTA AAA ATC 3' (containing a HindIII site; denoted SEQ ID NO:65). The resulting PCR product nfCP1₆₆₀ was submitted to nucleic acid sequence analysis to obtain a nucleic acid sequence of the coding strand, represented herein as SEQ ID NO:94. Translation of SEQ ID NO:94 indicated that nfCP1₆₉₆ encodes a protein of about 220 amino acids, called PfCP1₂₂₀, having SEQ ID NO:95. It is to be noted that this sequence analysis indicated that the stop codon was actually about 36 base pairs upstream from what had been predicted by SEQ ID NO:1; as such, the protein encoded by nfCP1₆₆₀ is about 12 amino acids shorter than would have been predicted by SEQ ID NO:1. The nucleic acid molecule nfCP1₆₆₀ contains the coding region for PfCP1₂₂₀.

Recombinant cell E. coli:pCro-nfCP1₆₆₀ is produced in the following manner. Nucleic acid molecule nfCP1₆₆₀ is digested with BamHI and HindIII restriction endonucleases, gel purified, and subcloned into expression vector lambdaP_(R) /T² ori/S10HIS-RSET-A9 (the production of which is described in Tripp et al, International PCT Publication No. WO 95/24198, published Sep. 14, 1995; see in particular, Example 7), that is digested with BamHI and HindIII and dephosphorylated. The resultant recombinant molecule, referred to herein as pCro-nfCP1₆₆₀, is transformed into E. coli BL-21 competent cells (available from Novagen, Madison, Wis.) to form recombinant cell E. coli:pCro-nfCP1₆₆₀. The recombinant cell is cultured as described in Example 20 of related PCT Publication No. WO 95/24198. About 1 ml of culture is collected prior to induction, and about 1 ml of culture is collected about 60 minutes following induction. These samples are then lysed in sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) loading buffer, resolved on a 14% Tris-glycine acrylamide gel and analyzed by immunoblot using a T7 (tag) antibody (available from Novagen).

Example 9

This example provides additional nucleic acid and deduced amino acid sequences of nucleic acid molecules encoding serine protease proteins of the present invention which are described herein and in the Examples section of related PCT Publication No.WO 96/11706.

A. A DNA probe labeled with ³² p comprising nucleotides from nfAP2₂₁₀₀ (described in Example 23 of related U.S. patent application Ser. No. 08/639,075, filed Apr. 24, 1996) was used to screen a bovine blood-fed whole flea cDNA library (described in Example 8 of related PCT Publication No.WO 96/11706) using standard hybridization techniques. A clone was isolated having about a 459-nucleotide insert, referred to herein as nfSP18₄₅₉.

A nucleic acid sequence of the composite nucleic acid molecule produced using nucleic acid sequence from nfSP18₅₃₄ and nfSP18₄₅₉ is referred to herein as nfSP18₇₇₉, having a nucleic acid sequence of the coding strand which is denoted herein as SEQ ID NO:9. Translation of SEQ ID NO:9 suggests that nucleic acid molecule nfSP18₇₇₅ encodes a non-full-length flea serine protease protein of about 228 amino acids, referred to herein as PfSP18₂₂₈, having amino acid sequence SEQ ID NO:10, assuming the first codon spans from about nucleotide 1 through about nucleotide 3 of SEQ ID NO:9 and the stop codon spans from about nucleotide 685 through about nucleotide 687 of SEQ ID NO:9. The complement of SEQ ID NO:9 is represented herein by SEQ ID NO:11. The coding region encoding PfSP18₂₂₈, is represented by nucleic acid molecule nfSP18₂₂₅, having a coding strand with the nucleic acid sequence represented by SEQ ID NO:12 and a complementary strand with nucleic acid sequence SEQ ID NO:14. The amino acid sequence of PfSP18₂₂₈ (i.e., SEQ ID NO:10) predicts that PfSP18₂₂₈ has an estimated molecular weight of about 25 kD and an estimated pI of about 9.09.

Comparison of nucleic acid sequence SEQ ID NO:9 with nucleic acid sequences reported in GenBank indicates that SEQ ID NO:9 showed the most homology, i.e., about 51% identity, between SEQ ID NO:9 and an Anopheles stephensi trypsin 1 gene. Comparison of amino acid sequence SEQ ID NO:10 (i.e., the amino acid sequence of PfSP18₂₂₈) with amino acid sequences reported in GenBank indicates that SEQ ID NO:10 showed the most homology, i.e., about 59% identity between SEQ ID NO:10 and Vespa crabro protein.

B. The remainder of flea serine protease nucleic molecule clone 24 (described in Example 2 was determined using primers designed from nfSP24₄₁₀ to amplify DNA from the bovine blood-fed whole flea cDNA library. Sense primer Flea 24F having the nucleotide sequence 5' GGA CAA ACT GTT CAT TGC AG 3' (denoted SEQ ID NO:46) was used in combination with the M13 universal primer in a first PCR reaction. Anti-sense primer Flea 24R having the nucleotide sequence 5' CCC TCA TTT GTC GTA ACT CC 3' (denoted SEQ ID NO:47) was used in combination with the M13 reverse primer in a second PCR reaction. The resulting PCR products were each gel purified and cloned into the TA Vector® System, and subjected to standard DNA sequencing techniques.

A composite nucleic acid sequence representing a flea serine protease coding region was deduced, referred to herein as nfSP24₁₀₈₉, was deduced and is denoted herein as SEQ ID NO:15. SEQ ID NO:78 is contained within the sequence of the nucleic acid molecule nfSP24₁₀₈₉. Translation of SEQ ID NO:15 suggests that nucleic acid molecule nfSP24₁₀₈₉ encodes a full-length flea serine protease protein of about 258 amino acids, referred to herein as PfSP24₂₅₈, having amino acid sequence SEQ ID NO:16, assuming an open reading frame in which the initiation codon spans from about nucleotide 33 through about nucleotide 35 of SEQ ID NO:15 and the termination codon spans from about nucleotide 807 through about nucleotide 809 of SEQ ID NO:15. The complement of SEQ ID NO:15 is represented herein by SEQ ID NO:17. The coding region encoding PfSP24₂₅₈, is represented by nucleic acid molecule nfSP24₇₇₄, having a coding strand with the nucleic acid sequence represented by SEQ ID NO:18 and a complementary strand with nucleic acid sequence SEQ ID NO:20. The proposed mature protein, denoted herein as PfSP24₂₃₇, contains about 237 amino acids which is represented herein as SEQ ID NO:22. The nucleic acid molecule encoding PfSP24₂₃₇ is denoted herein as nfSP24₇₁₁, which is represented by SEQ ID NO:21. The amino acid sequence of PfSP24₂₅₈ (i.e., SEQ ID NO:16) predicts that PfSP24₂₅₈ has an estimated molecular weight of about 28 kD and an estimated pI of about pI 6.70.

Comparison of nucleic acid sequence SEQ ID NO:15 with nucleic acid sequences reported in GenBank indicates that SEQ ID NO:15 showed the most homology, i.e., about 51% identity between SEQ ID NO:15 and an Anopheles stephensi trypsin 1 gene. Comparison of amino acid sequence SEQ ID NO:16 (i.e., the amino acid sequence of PfSP24₂₅₈) with amino acid sequences reported in GenBank indicates that SEQ ID NO:16 showed the most homology, i.e., about 59% identity between SEQ ID NO:16 and an Anopheles gambiae chymotrypsin II protein.

C. The remainder of flea serine protease nucleic molecule clone 32 (described in Example 8 was determined using primers designed from nfSP32₄₃₃ to amplify DNA from the cat-fed whole flea cDNA library. Sense primer Flea 32F having the nucleotide sequence 5' GGC TAG GTT AGT GGA TTC TGG 3' (denoted SEQ ID NO:48) was used in combination with the M13 universal primer in a first PCR reaction. Anti-sense primer Flea 32R having the nucleotide sequence 5' GCA AAT CAG TTC CAG AAT CCA CTA ACC 3' (denoted SEQ ID NO:49) was used in combination with the M13 reverse primer in a second PCR reaction. The resulting PCR products were each gel purified and cloned into the TA Vector® System, and subjected to standard DNA sequencing techniques.

A composite nucleic acid sequence representing a flea serine protease coding region was deduced, referred to herein as nfSP32₉₂₄, was deduced and is denoted herein as SEQ ID NO:23. SEQ ID NO:80 is contained within the sequence of the nucleic acid molecule nfSP32₉₂₄. Translation of SEQ ID NO:23 suggests that nucleic acid molecule nfSP32₉₂₄ encodes a full-length flea serine protease protein of about 268 amino acids, referred to herein as PfSP32₂₆₈, having amino acid sequence SEQ ID NO:24, assuming an open reading frame in which the initiation codon spans from about nucleotide 6 through about nucleotide 8 of SEQ ID NO:23 and the termination codon spans from about nucleotide 810 through about nucleotide 812 of SEQ ID NO:23. The complement of SEQ ID NO:23 is represented herein by SEQ ID NO:25. The coding region encoding PfSP32₂₆₈, is represented by nucleic acid molecule nfSP32₆₉₉, having a coding strand with the nucleic acid sequence represented by SEQ ID NO:26 and a complementary strand with nucleic acid sequence SEQ ID NO:28. The amino acid sequence of PfSP32₂₆₈ (i.e., SEQ ID NO:24) predicts that PfSP32₂₆₈ has an estimated molecular weight of about 28.6 kD and an estimated pI of about pI 7.36.

Comparison of nucleic acid sequence SEQ ID NO:23 with nucleic acid sequences reported in GenBank indicates that SEQ ID NO:23 showed the most homology, i.e., about 52% identity between SEQ ID NO:23 and a Fusarium oxysporum preprotrypsin gene. Comparison of amino acid sequence SEQ ID NO:24 (i.e., the amino acid sequence of PfSP32₂₆₈) with amino acid sequences reported in GenBank indicates that SEQ ID NO:24 showed the most homology, i.e., about 63% identity between SEQ ID NO:24 and a Bombyx mori vitellin-degrading protease precursor protein.

D. The remainder of flea serine protease nucleic molecule clone 33 was determined using primers designed from nfSP33₇₇₈ to amplify DNA from the flea mixed instar larvae cDNA library described above in Example 19. Sense primer Flea 33F having the nucleotide sequence 5' CAG GGC GCT CTG CAG AAC GCA AC 3' (denoted SEQ ID NO:50) was used in combination with the M13 universal primer in a first PCR reaction. Anti-sense primer Flea 33R having the nucleotide sequence 5' ATT CCT CGT GGT TCA GTC GCT C 3' (denoted SEQ ID NO:51) was used in combination with the M13 reverse primer in a second PCR reaction. The resulting PCR products were each gel purified and cloned into the TA Vector® System, and subjected to standard DNA sequencing techniques.

A composite nucleic acid sequence representing a flea serine protease coding region was deduced, referred to herein as nfSP33₁₈₉₄, was deduced and is denoted herein as SEQ ID NO:29. SEQ ID NO:84 and SEQ ID NO:82 are contained within the sequence of the nucleic acid molecule nfSP33₁₈₉₄. Translation of SEQ ID NO:29 suggests that nucleic acid molecule nfSP33₁₈₉₄ encodes a full-length flea serine protease protein of about 400 amino acids, referred to herein as PfSP33₄₀₀, having amino acid sequence SEQ ID NO:30, assuming an open reading frame in which the initiation codon spans from about nucleotide 335 through about nucleotide 337 of SEQ ID NO:29 and the termination codon spans from about nucleotide 1535 through about nucleotide 1537 of SEQ ID NO:29. The complement of SEQ ID NO:29 is represented herein by SEQ ID NO:31. The coding region encoding PfSP33₄₀₀, is represented by nucleic acid molecule nfSP33₁₂₀₀, having a coding strand with the nucleic acid sequence represented by SEQ ID NO:32 and a complementary strand with nucleic acid sequence SEQ ID NO:34. The proposed mature protein, denoted herein as PfSP33₂₄₂, contains about 242 amino acids which is represented herein as SEQ ID NO:36. The nucleic acid molecule encoding PfSP33₂₄₂ is denoted herein as nfSP33₇₂₆, which is represented by SEQ ID NO:35. The amino acid sequence of PfSP33₄₀₀ (i.e., SEQ ID NO:30) predicts that PfSP33₄₀₀ has an estimated molecular weight of about 44 kD and an estimated pI of about pI 7.59.

Comparison of nucleic acid sequence SEQ ID NO:29 with nucleic acid sequences reported in GenBank indicates that SEQ ID NO:29 showed the most homology, i.e., about 48% identity between SEQ ID NO:29 and a Drosophila melanogaster serine protease stubble gene. Comparison of amino acid sequence SEQ ID NO:30 (i.e., the amino acid sequence of PfSP33₄₀₀) with amino acid sequences reported in GenBank indicates that SEQ ID NO:30 showed the most homology, i.e., about 63% identity between SEQ ID NO:30 and a Drosophila melanogaster serine protease stubble protein.

Example 10

This example provides nucleic acid and deduced amino acid sequence of another nucleic acid molecule encoding a serine protease protein of the present invention.

A serine protease cDNA nucleic acid molecules has been isolated in a manner similar to that described in Example 8 of related PCT Publication No.WO 96/11706. The actual primers used in PCR amplification of the serine protease nucleic acid molecule from a cat blood-fed flea cDNA expression library (produced as described in Example 8 of related PCT Publication No.WO 96/11706) included cat-try #2 (SEQ ID NO:86) in combination with H57 primer (SEQ ID NO:99). The resultant PCR product was gel purified and cloned into the TA Vector™. A recombinant TA vector clone was isolated and subjected to nucleic acid sequencing.

A composite nucleic acid sequence of a flea serine protease nucleic molecule corresponding to flea clone 40, namely nfSP40₄₂₈ was deduced and is denoted herein as SEQ ID NO:37. Translation of SEQ ID NO:37 suggests that nucleic acid molecule nfSP40₄₂₈ encodes a non-full-length flea serine protease protein of about 142 amino acids, referred to herein as PfSP40₁₄₂, represented herein by SEQ ID NO:38. The complement of SEQ ID NO:37 is represented herein by SEQ ID NO:39.

The remainder of flea serine protease nucleic molecule clone 40 was determined using primers designed from nfSP40₄₂₈ to amplify DNA from the cat blood-fed whole flea cDNA library. Sense primer Flea 40F having the nucleotide sequence 5' GGC AAG TTT CGT TTC ACA ATA GG 3' (denoted SEQ ID NO:52) was used in combination with the M13 universal primer in a first PCR reaction. Anti-sense primer Flea 40R having the nucleotide sequence 5' TCC AAC CCT AAC TTT TAA ACC TTC 3' (denoted SEQ ID NO:53) was used in combination with the M13 reverse primer in a second PCR reaction. The resulting PCR products were each gel purified and cloned into the TA Vector® System, and subjected to standard DNA sequencing techniques.

A composite nucleic acid sequence representing a flea serine protease coding region was deduced, referred to herein as nfSP40₈₄₁, was deduced and is denoted herein as SEQ ID NO:40. SEQ ID NO:37 is contained within the sequence of the nucleic acid molecule nfSP40₈₄₁. Translation of SEQ ID NO:40 suggests that nucleic acid molecule nfSP40₈₄₁ encodes a non-full-length flea serine protease protein of about 242 amino acids, referred to herein as PfSP40₂₄₂, having amino acid sequence SEQ ID NO:41, assuming an open reading frame in which the first codon spans from about nucleotide 2 through about nucleotide 4 of SEQ ID NO:40 and the termination codon spans from about nucleotide 728 through about nucleotide 730 of SEQ ID NO:40. The complement of SEQ ID NO:40 is represented herein by SEQ ID NO:42. The coding region encoding PfSP40₂₄₂, is represented by nucleic acid molecule nfSP40₇₁₇, having a coding strand with the nucleic acid sequence represented by SEQ ID NO:43 and a complementary strand with nucleic acid sequence SEQ ID NO:45. The amino acid sequence of PfSP40₂₄₂ (i.e., SEQ ID NO:41) predicts that PfSP40₂₄₂ has an estimated molecular weight of about 26 kD and an estimated pI of about pI 6.5.

Comparison of nucleic acid sequence SEQ ID NO:40 with nucleic acid sequences reported in GenBank indicates that SEQ ID NO:40 showed the most homology, i.e., about 57% identity between SEQ ID NO:40 and a Dermatophagoides pteronyssinus Der P3 allergen gene. Comparison of amino acid sequence SEQ ID NO:41 (i.e., the amino acid sequence of PfSP40₂₄₂) with amino acid sequences reported in GenBank indicates that SEQ ID NO:41 showed the most homology, i.e., about 40% identity between SEQ ID NO:41 and a Bombyx mori vitellin-degrading protease precursor protein.

Example 11

This Example demonstrates the production of serine protease proteins of the present invention in E. coli cells.

A. Flea serine protease protein PfSP24₂₅₈ was produced in the following manner. An about 714 bp nucleic acid molecule, referred to herein as nfSP24₇₁₄ (designed to encode an apparently mature serine protease protein) was PCR amplified from nfSP24₁₀₈₉ using sense primer Flea 24 EF having the nucleotide sequence 5' CAC AGG ATC CAA TAA TTT GTG GTC AAA ATG C 3' (containing a BamHI-site; denoted SEQ ID NO:54) and anti-sense primer Flea 24 ER having the nucleotide sequence 5' AAA AAG AAA GCT TCT TTA ATT TTC TGA CAT TGT CGT G 3' (containing a HindIII; denoted SEQ ID NO:55). The resulting PCR product nfSP24₇₁₄ was digested with BamHI and HindIII restriction endonucleases, gel purified, and subcloned into expression vector lambdaP_(R) /T² ori/S10HIS-RSET-A9, that had been digested with BamHI and HindIII and dephosphorylated. The resultant recombinant molecule, referred to herein as pCro-nfSP24₇₁₄, was transformed into E. coli BL-21 competent cells (available from Novagen, Madison, Wis.) to form recombinant cell E. coli:pCro-nfSP24₇₁₄. The recombinant cell was cultured as described in Example 20 of related PCT Publication No.WO 95/24198. About 1 ml of culture was collected prior to induction, and about 1 ml of culture was collected about 60 minutes following induction. These samples were then lysed in sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) loading buffer and resolved on a 14% Tris-glycine acrylamide gel. Immunoblot analysis of the proteins using a T7 (tag) antibody (available from Novagen) showed expression of an about 36 kD protein in the induced sample but not in the uninduced sample.

B. Flea serine protease protein PfSP32₂₆₈ was produced in the following manner. An about 698 bp nucleic acid molecule, referred to herein as nfSP32₆₉₈ (designed to encode an apparently mature serine protease protein) was PCR amplified from nfSP32₉₃₃ using sense primer Flea 32 EF having the nucleotide sequence 5' GCG GGA TCC TAT TGT GGG TGG TGA AGC AGT G 3' (containing a BamHI-site; denoted SEQ ID NO:56) and anti-sense primer Flea 32 ER having the nucleotide sequence 5' GAC GGT ACC ATG TAT AAA ATA ATA TTA AAC TCC GG 3' (containing a KpnI; denoted SEQ ID NO:57). The resulting PCR product nfSP32₆₉₈ was digested with BamHI and KpnI restriction endonucleases, gel purified, and subcloned into expression vector 1 pTrcHisB (available from Invitrogen Corp., San Diego, Calif.), that had been digested with BamHI and KpnI and dephosphorylated. The resultant recombinant molecule, referred to herein as pTrc-nfSP32₆₉₈, was transformed into E. coli BL-21 competent cells to form recombinant cell E. coli:pTrc-nfSP32₆₉₈. The recombinant cell was cultured and protein production resolved by SDS-PAGE as described above in Section A. Immunoblot analysis of the proteins using a T7 antibody showed expression of an about 38 kD protein in the induced sample but not in the uninduced sample.

C. Flea serine protease protein PfSP33₄₀₀ was produced in the following manner. An about 1200 bp nucleic acid molecule, referred to herein as nfSP33₁₂₀₀ (designed to encode an apparently mature serine protease protein) was PCR amplified from nfSP33₁₈₉₄ using sense primer Flea 33 EF having the nucleotide sequence 5' CCG GGA TCC TAT GTT AGC GAT CGT CCC GTC AAA C 3' (containing a BamHI-site; denoted SEQ ID NO:58) and anti-sense primer Flea 33 ER having the nucleotide sequence 5' CCG GAA TTC TTA TCC CAT TAC TTT GTC GAT CC 3' (containing a EcoRI; denoted SEQ ID NO:59). The resulting PCR product nfSP33₁₂₀₀ was digested with BamHI and EcoRI restriction endonucleases, gel purified, and subcloned into expression vector lambdaP_(R) /T² ori/S10HIS-RSET-A9, that had been digested with BamHI and EcoRI and dephosphorylated. The resultant recombinant molecule, referred to herein as pCro-nfSP33₁₂₀₀, was transformed into E. coli BL-21 competent cells to form recombinant cell E. coli:pCro-nfSP33₁₂₀₀. The recombinant cell was cultured using the method described above in Section A.

D. Flea serine protease protein PfSP40₂₄₂ was produced in the following manner. An about 716 bp nucleic acid molecule, referred to herein as nfSP40₇₁₆ (designed to encode an apparently mature serine protease protein) was PCR amplified from nfSP40₈₄₁ using sense primer Flea 40 EF having the nucleotide sequence 5' GCG GGA TCC AAT AGT AGG AGG TGA AGA TGT AG 3' (containing a BamHI-site; denoted SEQ ID NO:60) and anti-sense primer Flea 40 ER having the nucleotide sequence 5' CCG GAA TTC TTC TAA CAA ATT TTA TTT GAT TCC TGC 3' (containing a EcoRI; denoted SEQ ID NO:61). The resulting PCR product nfSP40₇₁₆ was digested with BamHI and EcoRI restriction endonucleases, gel purified, and subcloned into expression vector lambdaP_(R) /T² ori/S10HIS-RSET-A9, that had been digested with BamHI and EcoRI and dephosphorylated. The resultant recombinant molecule, referred to herein as pCro-nfSP40₇₁₆, was transformed into E. coli BL-21 competent cells to form recombinant cell E. coli:pCro-nfSP40₇₁₆. The recombinant cell was cultured and protein production resolved using the methods described above in Section A. Immunoblot analysis of the proteins using a T7 antibody showed expression of an about 38 kD protein in the induced sample but not in the uninduced sample.

Example 12

This Example demonstrates the production of another serine protease protein of the present invention in E. coli cells.

A. Flea serine protease protein PfSP28₂₃₇ was produced in the following manner. An about 711 bp nucleic acid molecule, referred to herein as nfSP28₇₁₁ (designed to encode an apparently mature serine protease protein) was PCR amplified from nfSP28₉₂₃ using sense primer Flea 28 F having the nucleotide sequence 5' GGA TCC AAT CGT TGG AGG TGA AGA TG 3' (containing a BamHI-site shown in bold; denoted SEQ ID NO:62) and anti-sense primer Flea 28 R having the nucleotide sequence 5' GAA TTC GAA ATC CAC TTA AAC ATT AGC 3' (containing a EcoRI shown in bold; denoted SEQ ID NO:63). The resulting PCR product nfSP28₇₁₁ was digested with BamHI and EcoRI restriction endonucleases, gel purified, and subcloned into expression vector lambdaP_(R) /T² ori/S10HIS-RSET-A9, that had been digested with BamHI and XbaI and dephosphorylated. The resultant recombinant molecule, referred to herein as pCro-nfSP28₇₁₁, was transformed into E. coli BL-21 competent cells (available from Novagen, Madison, Wis.) to form recombinant cell E. coli:pCro-nfSP28₇₁₁. The recombinant cell was cultured and protein production resolved using the methods described above in Example 21. Immunoblot analysis of the proteins using a T7 antibody showed expression of an about 38 kD protein in the induced sample but not in the uninduced sample. Immunoblot analysis using a rabbit anti-flea midgut protease polyclonal antibody (the production of which is described in Example 14 of related PCT Publication NO.WO 95/24198) identified an about 38 kD protein in the induced sample.

Example 13

This Example demonstrates the production of another serine protease protein of the present invention in eukaryotic cells.

Recombinant molecule pBv-nfSP28₇₉₂, containing a flea serine protease nucleic acid molecule spanning nucleotides from about 11 through about 802 of SEQ ID NO:66, operatively linked to baculovirus polyhedron transcription control sequences were produced in the following manner. In order to subclone a flea serine protease nucleic acid molecule into a baculovirus expression vector, a flea serine protease nucleic acid molecule-containing fragment was PCR amplified from nfSP28₉₂₃. A PCR fragment of 792 nucleotides, named nfSP28,₇₉₂, was amplified from nfSP28₉₂₃ using a sense primer Flea 28 F3 having the nucleic acid sequence 5'-GCG GGA TTC TAT AAA TAT GAA ACT TTT GGT AGT TTT TGC -3' (SEQ ID NO:62; BamHI site shown in bold) and an antisense primer Flea 28 R3 having the nucleic acid sequence 5'-GCT CTA GAC CAC TTA AAC ATT AGC ATA TTT TTC- 3'(SEQ ID NO:63; XbaI site shown in bold). The N-terminal primer was designed from the pol h sequence of baculovirus with modifications to enhance expression in the baculovirus system.

In order to produce a baculovirus recombinant molecule capable of directing the production of PfSP28₂₆₄, the about 792 base pair PCR product (referred to as Bv-nfSP28₇₉₂) was digested with BamHI and XbaI and subcloned into BamHI and XbaI digested to produce the recombinant molecule referred to herein as pVL-nfSP28₇₉₂.

The resultant recombinant molecule,pVL-nfSP28₇₉₂, was verified for proper insert orientation by restriction mapping. The recombinant molecule was co-transfected with a linear Baculogold baculovirus DNA (available from Pharmingen) into S. frugiperda Sf9 cells (available from InVitrogen) to form the recombinant cells denoted S. frugiperda:pVL-nfSP28₇₉₂. S. frugiperda:pVL-nfSP28₇₉₂ was cultured in order to produce a flea serine protease protein PfSP28₂₆₄.

Immunoblots of supernatants from cultures of S. frugiperda:pVL-nfSP28₇₉₂ cells producing the flea serine protease protein PfSP28₂₆₄ was performed using a cat anti-fSPFlea 28 polyclonal antibody which was produced as follows. Recombinant Flea 28 protein (referred to herein as rSPFlea 28 protein) produced in E. coli described above in Example 12 was used to immunize cats. The rSPFlea 28 protein was diluted to a concentration of about 1 mg/ml in PBS and emulsified in an equal volume of TiterMax research adjuvant (available from CytRx Corp., Norcross, Ga.). A series of cats were immunized each with about 50 μg of rSPFlea 28 protein in adjuvant by subcutaneous injection. A second injection of the same dose of rSPFlea 28 protein in adjuvant was administered 32 days later. Blood samples were obtained prior to immunization (pre-bleed), 32 days and 47 days after the initial immunization. Sera samples from the pre-immunization and Day 47 bleeds were used for subsequent immunoblot experiments. The latter is referred to as anti-fSPFlea 28 polyclonal antibody.

Analysis of the immunoblots identified an about 33 kD protein and an about 36 kD protein.

Example 14

This example describes the production of peptides from the 31 kD flea midgut serine protease and the generation of internal sequence data.

Midguts from about 30,000 cat blood-fed fleas were dissected as described in U.S. Pat. No. 5,356,622, ibid. in gut dissection buffer (50 mM Tris 8.0, 100 mM CaCl₂) . The guts (in three batches of about 10,000 each) were disrupted by a freeze-thaw cycle, followed by sonication. The resulting extracts were clarified by centrifugation for 20 minutes at 3050 rpm in a swinging bucket centrifuge at 4° C. The supernatants were recovered, and adjusted to 400 mM NaCl in preparation for benzamidine column chromatography.

For each batch, gut supernatants were loaded into a 5 ml disposable column containing p-aminobenzamidine cross-linked to Sepharose beads (available from Sigma, St. Louis, Mo.), previously equilibrated in benzamidine column buffer (50 mM Tris, pH 8.0, 100 mM CaCl₂, 400 mM NaCl) and incubated with rocking overnight at 4° C. Unbound protein was slowly washed off the column using benzamidine column buffer until no protein was detectable by Bradford Assay (available from Bio-Rad Laboratories, Hercules, Calif.).

Proteases bound to the benzamidine column were eluted using 4 ml benzamidine column buffer supplemented with 100 mM p-aminobenzamidine (brought to pH 8.0 with NaOH). Residual bound proteases were washed off with about 21 ml of additional benzamidine column buffer. The recovered proteases were then concentrated to a volume of about 2 ml using a Ultrafree 20 10-kD centrifugal concentrator (available from Millipore, Bedford, Mass.). After concentration, the protease pools from the 3 preparations were combined for a total of about 30,000 gut equivalents in about 6 ml. Protein concentration was measured by Bradford assay and found to be about 0.5 mg/ml.

About 150 μg of the isolated protease pool was resolved by polyacrylamide gel electrophoresis (PAGE) on a preparative-well 14% Tris-glycine gel (available from Novex, San Diego, Calif.). After electrophoresis, the proteins in the gel were visualized by staining for about 30 minutes in Coomassie brilliant blue stain (0.1% (w/v) Coomassie blue R, 40% (v/v) methanol, 10% (v/v) acetic acid) and destaining for about 2.5 hours in 50% (v/v) methanol. The band corresponding to the 31-kD protease was excised with a razor blade. The protein was electroeluted, concentrated, and partially digested for 24 hours with cyanogen bromide (CNBr) (Silver, et al., 1995, J. Biol. Chem., 270, 13010-13016), except that a small amount of acetic acid was added to the sample after electroelution and concentration to lower the sample pH and therefore reduce autodigestion by the 31-kD protease. CNBr is known to cleave after methionine (M) residues under the conditions used for this digestion. After CNBr digestion, the peptides in the sample were resolved by PAGE on an 18% Tris-glycine gel. After electrophoresis, the separated protease peptides were electroblotted onto a PVDF membrane using a CAPS buffer (10 mM CAPS pH 11, 0.5 mM DTT, 10% (v/v) methanol). The membrane was stained with Coomassie Brilliant Blue and destained with 50% (v/v) methanol. Three stained peptide bands were identified having apparent molecular weights of about 14 kD, 21 kD, and 22 kD, respectively. The portions of the membrane containing the 21 kD and 22 kD bands were excised separately. Peptides contained in each membrane segment were subjected to N-terminal amino acid sequencing using a 473A Protein Sequencer (available from Applied Biosystems, Foster City, Calif.) using standard techniques.

Although the results from the automated sequencing were difficult to interpret due to overlapping sequences, analysis of the chromatograms indicated the N-terminal amino acid sequence of the 21-kD peptide to be H/R-V/P-G/A/S-Y/G-E/N-D/K-V/R-D/A-D-Y- D-F-D/P-V-A, denoted herein as SEQ ID NO:70 and the N-terminal amino acid sequence of the 22-kD peptide to be I/Q-V-G-Y/G-E/N/T-D/M/P-V-K/D-I-N/S-M/T/N-F/C herein denoted as SEQ ID NO:71. The N-terminal amino acid sequence of the intact 31-kD protease is either I-V-G-G- E-D-V-D-I-S-T-C-G-W-C (SEQ ID NO:59, as disclosed in Example 34 in co-pending U.S. patent application Ser. No. 08/639,075), or IVGGEDVDIST(C)GWQI(S)FQ(S)ENLHF(C)GG(S)IIAPK (SEQ ID NO:69, as disclosed in Example 35 in co-pending U.S. patent application Ser. No. 08/639,075). These sequences vary at residue 15 in that SEQ ID NO:68 contains a cysteine and SEQ ID NO:69 contains a glutamine. These sequences can be identified in the sequences of both the 21-kD (SEQ ID NO:70) and 22-kD (SEQ ID NO:71) peptides, though it is much stronger in the 22-kD peptide, leading to the conclusion that the SEQ ID NO:71 represents the N-terminus of the 31-kD protease. If this sequence is subtracted from the sequence for the 21-kD (SEQ ID NO:70) peptide, then the resulting sequence for the 21-kD peptide is H/R-P-A/S-Y-N-K-R-A-D-Y-D-F-D-V-A, denoted herein as SEQ ID NO:72. This sequence of amino acids aligns with a stretch of deduced amino acids from about residue 107 to residue 121 immediately following a methionine residue in SEQ ID NO:67. These data confirm that the clone represented by nfSP28₉₂₃ (SEQ ID NO:66, as disclosed in Example 36 in co-pending U.S. patent application Ser. No. 08/639,075) indeed encodes the 31-kD protease.

Example 15

This example demonstrates that a 31-kD flea midgut serine protease contained in a formulation is able to proteolyze cat immunoglobulin G, A, and M proteins as well as bovine, dog, human, and rabbit immunoglobulin G proteins.

The 31-kD flea midgut serine protease was purified from cat blood-fed fleas as follows. Cat blood-fed flea midgut extracts were prepared and selected on a benzamidine column as described above in Example 25. The benzamidine eluate was then further purified as described in Example 35 of co-pending U.S. patent application Ser. No. 08/639,075 by PolyCAT A cation exchange chromatography (available from PolyLC, Inc., Columbia, Md.) to isolate a protein band which migrated at about 31 kD on a silver stained SDS-PAGE gel.

A. The ability of the cat blood-fed 31-kD flea midgut serine protease to degrade immunoglobulin was demonstrated by measuring digestion of immunoglobulin heavy chain using a method similar to that described in Example 35 of co-pending U.S. patent application Ser. No. 08/639,075. Specifically, 1 μg samples of cat IgG, cat IgA, and cat IgM substrates (available from Bethyl Laboratories, Inc., Montgomery, Tex.) were incubated with 500 cat blood-fed flea midgut equivalents of purified 31-kD flea midgut serine protease in a total volume of 27 μl 0.1M Tris-HCl pH 8.0 at 37° C. for 18 hours. The reaction mixtures were resolved by 14% Tris-glycine SDS-PAGE and the gel was silver stained using standard methods. The total disappearance (compared with control samples lacking addition of the purified 31-kD protein) of bands migrating at about 50, 60, and 80 kD on the silver stained gel in the lanes containing 31-kD protease-treated cat IgG, IgA, and IgM, respectively, indicated that the 31-kD flea midgut serine protease degraded the heavy chains of the various cat immunoglobulin isotypes.

B. The ability of the cat blood-fed 31-kD flea midgut serine protease to degrade IgG from several species was demonstrated by incubating 1 μg samples of purified cat or bovine IgG (purified from cat and bovine blood on Protein A Sepharose), or of purified dog, rabbit, or human IgG (each available from Sigma Chemical Co.) with 500 cat blood-fed flea midgut equivalents of the purified 31-kD flea midgut serine protease in a total volume of 27 μl 0.1M Tris-HCl pH 8.0 at 37° C. for 18 hours. The reaction mixtures were resolved by 14% Tris-glycine SDS-PAGE and the gel was silver stained using standard methods. The total disappearance (compared with control samples lacking addition of the purified 31-kD protein) of bands migrating at about 50-55-kD on the silver stained gel in the lanes containing the 31-kD protease treated cat, bovine, dog, rabbit, and human IgG heavy chains, indicated that the 31-kD flea midgut serine protease can degrade IgG from various mammalian species.

Example 16

This example describes the ability of a 31-kD flea midgut serine protease contained in a formulation to proteolyze cat immunoglobulin G at a specific site.

The 31-kD flea midgut serine protease was purified from cat blood-fed flea midgut extracts as described above in Examples 14 and 15

To investigate cleavage site specificity of the purified 31-kD flea midgut serine protease, 10 μg of cat immunoglobulin G purified from cat blood on Protein A sepharose was incubated with 200 cat blood-fed flea midgut equivalents of purified 31-kD flea midgut serine protease in a total volume of 100 μl 0.2 M Tris-HCl pH 8.0 at 37° C. for 18 hours. The reaction mixture was resolved by 14% Tris-glycine SDS-PAGE, blotted onto a PVDF membrane, stained with Coomassie R-250 and destained according to standard procedures. A band of about 33 kD was excised and subjected to N-terminal amino acid sequencing using techniques known to those skilled in the art. A partial N-terminal amino acid sequence of about 28 amino acids was determined and is represented herein as SEQ ID NO: 73: X-P-P-P-E-M-L-G-G-P-S-I-F-I-F-P-P-K-P-K-D-D-L-L-I-K-R-K. A GenBank homology search using SEQ ID NO:73 revealed most homology to Oryctolagus caniculus gamma H-chain constant region 2, there being about 71% identity over the 28 amino acids. Further alignments of SEQ ID NO:73 with sheep, rat, rabbit, monkey, bovine, and human IgG amino acid sequences indicated that the purified cat blood-fed 31-kD flea midgut serine protease cleaved the cat IgG heavy chain just before the predicted C-terminal end of the IgG hinge region. The predicted first amino acid cysteine and the second amino acid proline occur within the predicted hinge region while the remaining 26 amino acids of SEQ ID NO:73, starting with the third amino acid proline, occur within the predicted constant heavy chain-2 region.

The further investigate the cleavage site specificity of the purified 31-kD flea midgut serine protease for cat IgG, the cleavage site was compared to that of a known protease, papain, as follows. Cat immunoglobulin G (100 mg), purified from cat blood on Protein A sepharose, was incubated with 1 mg papain in 100 mM sodium acetate pH 5.5, 50 mM cysteine, 1 mM EDTA in a final volume of 150 μl at room temperature for 4.5 hours. The reaction mixture was resolved on a 14% Tris-glycine SDS-PAGE gel, blotted onto PVDF membrane, stained with Coomassie R-250 and destained according to standard procedures. A band of about 33 kD was excised and subjected to N-terminal amino acid sequencing using techniques known to those skilled in the art. A partial N-terminal amino acid sequence of about 25 amino acids was deduced and is represented herein as SEQ ID NO: 96: X-P-P-P-E-M-L-G-G-P-S-I-F-I-F-P-P-K-K-K-D-D-L-L-I. This sequence was nearly identical to the one obtained from a 33-kD cat IgG cleavage product generated by the purified 31-kD flea midgut serine protease, the only difference being the substitution of a lysine (SEQ ID NO:73) for a proline (SEQ ID NO:96) at amino acid 19.

Example 17

This Example demonstrates the kinetics of cat IgG degrading activity in the midguts of fleas fed on live cats.

A. To determine the kinetics of cat IgG degradation in the guts of continuously feeding fleas, female fleas contained in chambers were fed on seven separate cats (i.e. one chamber per cat) as described in Example 21 of co-pending U.S. patent application Ser. No. 08/639,075. Flea chambers were removed for dissections at timepoints of 15 min., 30 min., 1 hr., 2 hr., 4 hr., 6 hr., 8 hr., and 17 hr.. After feeding on the cats, the fleas, midguts were removed as described in U.S. Pat. No. 5,356,622, ibid., homogenized by freeze-fracture and sonicated in a Tris buffer comprising 50 mM Tris, pH 8.0 and 100 mM CaCl₂. The extracts were centrifuged at about 14,000×g for 20 min. and the soluble material recovered. The soluble material was then diluted to a final concentration of about 1.2 midgut equivalents per microliter (μl) of Tris buffer. The proteins contained in 1 midgut equivalent of each timepoint were then resolved by SDS-PAGE under reducing conditions, and the proteins visualized by silver staining. The results indicated that IgG heavy chain levels were significantly lower at the 17 hour time point than in the 8 hour and earlier time points, and that light chain levels were reduced but not to the same extent as the heavy chain. The proteins contained in 5 gut equivalents of each timepoint were then resolved by SDS-PAGE gel under reducing conditions and were subjected to western blot analysis using alkaline phosphatase labeled goat anti-cat IgG (heavy plus light chain) antibody (available from Kirkegaard and Perry Laboratories, Gaithersburg, Md.). The results indicated that cat IgG heavy chain was present in the midguts of continuously feeding fleas for at least 8 hours, but was not detected in the midguts of fleas allowed to feed continuously on a cat for 17 hours. Light chain was visible in all samples, though the amount visible in the 17 hour sample was significantly less than that visible in the 8 hour sample. These results suggest that even when fleas are continuously feeding on a cat, the levels of IgG-degrading proteases induced in the flea midguts at a time point of 17 hours is sufficient to degrade all detectable cat IgG ingested. These results suggest that when fleas are continuously feeding on a cat, the levels of IgG-degrading proteases induced in the flea midguts are not sufficient to degrade all detectable cat IgG ingested for at least 8 hours.

B. To determine the kinetics of cat IgG degradation in the guts of fleas fed for a specified time then removed from the cat, fleas (in chambers) were fed on cats as in Section A for periods of either 1 hour or 24 hours. Following the 1 or 24-hour feeding periods, the flea chambers were removed and placed in a 28° C., 75% relative humidity growth incubator. Fleas were subjected to dissection at time points of 0, 1, 2, 4, and 8 hr. following removal from the cats. Midguts were homogenized, and the midgut contents were examined by silver stained SDS-PAGE and immunoblot analysis, as described in Section A. The fleas fed for 1 hour had high molecular weight proteins, including the heavy chain and light chain of cat IgG detectable in their midguts at the 0 and 1 hour dissection timepoints, while the flea midguts evaluated at time points of 2 hours or greater had no detectable IgG heavy chain bands. The results showed that when fleas were fed on a cat and then removed, they degraded the ingested cat IgG heavy chain nearly completely within 2 hours. The fleas fed on cats for 24 hours had no detectable IgG heavy or light chain proteins in midgut extracts at any of the timepoints. These results suggest that when no new cat IgG is ingested, as is the case when the fleas are removed from feeding, that the IgG-degrading proteases in the flea midgut fully degraded all cat IgG heavy chain in less than two hours.

    __________________________________________________________________________     #             SEQUENCE LISTING                                                   - -  - - (1) GENERAL INFORMATION:                                              - -    (iii) NUMBER OF SEQUENCES: 103                                          - -  - - (2) INFORMATION FOR SEQ ID NO:1:                                      - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 1109 base - #pairs                                                 (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: cDNA                                               - -     (ix) FEATURE:                                                                   (A) NAME/KEY: CDS                                                              (B) LOCATION: 126..1106                                               - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:1:                                - - TACAATTAGG GTGACGATCT TTTAGACAAG CGAAATTGAT AACGAAGTTT TC -              #GAAGTCGG     60                                                                  - - ATGAAGTAAA AACCTTGCGT TGGTTTCCCC GGTCCCAGGA TCAGGAACAG TT -             #GCACTTTA    120                                                                  - - CCCCA ATG AGG GAA TTC GTG CAT CCC CAT TTT - #ACC GAA CAT ATT GAT             167                                                                              Met Arg Glu Phe Val His Pro - #His Phe Thr Glu His Ile Asp                       1          - #     5             - #     10                             - - GAA GAA TTC CAC CGA TTC ATC AAT AAA CAC GG - #A AAA ATT TAT AAT AAA           215                                                                        Glu Glu Phe His Arg Phe Ile Asn Lys His Gl - #y Lys Ile Tyr Asn Lys             15                 - # 20                 - # 25                 - # 30        - - AAT GAA GAA CAT CAT TTC CGC AAA GAA ATT TT - #C AGA CTA AAC TTG AGG           263                                                                        Asn Glu Glu His His Phe Arg Lys Glu Ile Ph - #e Arg Leu Asn Leu Arg                             35 - #                 40 - #                 45               - - TAC ATT TTT TCT AAG AAT CGT GCA AAT TTG GG - #A TAC ACT TTG ACT GTT           311                                                                        Tyr Ile Phe Ser Lys Asn Arg Ala Asn Leu Gl - #y Tyr Thr Leu Thr Val                         50     - #             55     - #             60                   - - AAC CAT TTG GCT GAT CGT ACT GAA GCT GAA CT - #T AAG GCT TTG AGA GGA           359                                                                        Asn His Leu Ala Asp Arg Thr Glu Ala Glu Le - #u Lys Ala Leu Arg Gly                     65         - #         70         - #         75                       - - CAC AGA CCT TCC TCC GGT TAT AAT GGC GGT TT - #A CCC TTT CCT CAC AAT           407                                                                        His Arg Pro Ser Ser Gly Tyr Asn Gly Gly Le - #u Pro Phe Pro His Asn                 80             - #     85             - #     90                           - - ACC ACC AAG GAA GCA AGA AAT TTA CCA GAT TC - #T TTC GAC TGG CGA ATT           455                                                                        Thr Thr Lys Glu Ala Arg Asn Leu Pro Asp Se - #r Phe Asp Trp Arg Ile             95                 - #100                 - #105                 - #110        - - TAT GGA GCT GTT ACT CCA GTT AAA GAT CAA TC - #T GTT TGT GGT TCC TGC           503                                                                        Tyr Gly Ala Val Thr Pro Val Lys Asp Gln Se - #r Val Cys Gly Ser Cys                            115  - #               120  - #               125               - - TGG TCT TTC GGA ACA ATT GGA GCA ATC GAA GG - #T GCA TAT TTC TTG AAA           551                                                                        Trp Ser Phe Gly Thr Ile Gly Ala Ile Glu Gl - #y Ala Tyr Phe Leu Lys                        130      - #           135      - #           140                   - - AAC GGC GGT AAT CTT GTA CGA TTG TCT CAA CA - #G GCT TTG ATT GAT TGT           599                                                                        Asn Gly Gly Asn Leu Val Arg Leu Ser Gln Gl - #n Ala Leu Ile Asp Cys                    145          - #       150          - #       155                       - - TCT TGG GGA TAT GGA AAT AAT GGT TGC GAC GG - #T GGC GAA GAT TTC CGC           647                                                                        Ser Trp Gly Tyr Gly Asn Asn Gly Cys Asp Gl - #y Gly Glu Asp Phe Arg                160              - #   165              - #   170                           - - GCC TAC CAA TGG ATG ATG AAA CAT GGA GGA AT - #C CCT ACT GAA GAG GAT           695                                                                        Ala Tyr Gln Trp Met Met Lys His Gly Gly Il - #e Pro Thr Glu Glu Asp            175                 1 - #80                 1 - #85                 1 -       #90                                                                               - - TAT GGT GGT TAC TTG GGA CAA GAT GGT TAC TG - #C CAT GTC AAC AAC         GTT      743                                                                     Tyr Gly Gly Tyr Leu Gly Gln Asp Gly Tyr Cy - #s His Val Asn Asn Val                           195  - #               200  - #               205               - - ACT TTA GTT GCT CCC ATC ACA GGA TAT GTC AA - #C GTA ACT CGT AAC GAT           791                                                                        Thr Leu Val Ala Pro Ile Thr Gly Tyr Val As - #n Val Thr Arg Asn Asp                        210      - #           215      - #           220                   - - GTT GAC GCT ATG AAG GTT GCC CTT CTT AAA CA - #C GGT CCG ATT TCG GTG           839                                                                        Val Asp Ala Met Lys Val Ala Leu Leu Lys Hi - #s Gly Pro Ile Ser Val                    225          - #       230          - #       235                       - - GCC ATT GAC GCA TCA CAC AAA ACA TCC AGT TT - #T TAC TCC AAC GGC GTT           887                                                                        Ala Ile Asp Ala Ser His Lys Thr Ser Ser Ph - #e Tyr Ser Asn Gly Val                240              - #   245              - #   250                           - - TAC TAC CAA CCG AAA TGT GGC AAT AAA AGA GG - #A CAA TTA GAC CAC GCC           935                                                                        Tyr Tyr Gln Pro Lys Cys Gly Asn Lys Arg Gl - #y Gln Leu Asp His Ala            255                 2 - #60                 2 - #65                 2 -       #70                                                                               - - GTA TTA GTA GTC GGT TAT GGT GAA ATC AAC AG - #C GAA CCT TAC TGG         TTG      983                                                                     Val Leu Val Val Gly Tyr Gly Glu Ile Asn Se - #r Glu Pro Tyr Trp Leu                           275  - #               280  - #               285               - - GTC AAG AAT CCT GGT CAA TTG TGG GGA AAC AA - #T GTT ATA TTT GAT GTC          1031                                                                        Val Lys Asn Pro Gly Gln Leu Trp Gly Asn As - #n Val Ile Phe Asp Val                        290      - #           295      - #           300                   - - GCC AAA AAT AAT AAT GCG GAT TTG ACG ATC AC - #T TAT TTA CTA TGT ACT          1079                                                                        Ala Lys Asn Asn Asn Ala Asp Leu Thr Ile Th - #r Tyr Leu Leu Cys Thr                    305          - #       310          - #       315                       - - ACT TTT AAA ATT GAT TTT TAC GGC ATG TAA  - #                  - #              1109                                                                      Thr Phe Lys Ile Asp Phe Tyr Gly Met                                                320              - #   325                                                  - -  - - (2) INFORMATION FOR SEQ ID NO:2:                                      - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 327 amino - #acids                                                 (B) TYPE: amino acid                                                           (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:2:                                - - Met Arg Glu Phe Val His Pro His Phe Thr Gl - #u His Ile Asp Glu Glu         1               5 - #                 10 - #                 15               - - Phe His Arg Phe Ile Asn Lys His Gly Lys Il - #e Tyr Asn Lys Asn Glu                    20     - #             25     - #             30                   - - Glu His His Phe Arg Lys Glu Ile Phe Arg Le - #u Asn Leu Arg Tyr Ile                35         - #         40         - #         45                       - - Phe Ser Lys Asn Arg Ala Asn Leu Gly Tyr Th - #r Leu Thr Val Asn His            50             - #     55             - #     60                           - - Leu Ala Asp Arg Thr Glu Ala Glu Leu Lys Al - #a Leu Arg Gly His Arg        65                 - # 70                 - # 75                 - # 80        - - Pro Ser Ser Gly Tyr Asn Gly Gly Leu Pro Ph - #e Pro His Asn Thr Thr                        85 - #                 90 - #                 95               - - Lys Glu Ala Arg Asn Leu Pro Asp Ser Phe As - #p Trp Arg Ile Tyr Gly                   100      - #           105      - #           110                   - - Ala Val Thr Pro Val Lys Asp Gln Ser Val Cy - #s Gly Ser Cys Trp Ser               115          - #       120          - #       125                       - - Phe Gly Thr Ile Gly Ala Ile Glu Gly Ala Ty - #r Phe Leu Lys Asn Gly           130              - #   135              - #   140                           - - Gly Asn Leu Val Arg Leu Ser Gln Gln Ala Le - #u Ile Asp Cys Ser Trp       145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - Gly Tyr Gly Asn Asn Gly Cys Asp Gly Gly Gl - #u Asp Phe Arg Ala         Tyr                                                                                              165  - #               170  - #               175              - - Gln Trp Met Met Lys His Gly Gly Ile Pro Th - #r Glu Glu Asp Tyr Gly                   180      - #           185      - #           190                   - - Gly Tyr Leu Gly Gln Asp Gly Tyr Cys His Va - #l Asn Asn Val Thr Leu               195          - #       200          - #       205                       - - Val Ala Pro Ile Thr Gly Tyr Val Asn Val Th - #r Arg Asn Asp Val Asp           210              - #   215              - #   220                           - - Ala Met Lys Val Ala Leu Leu Lys His Gly Pr - #o Ile Ser Val Ala Ile       225                 2 - #30                 2 - #35                 2 -       #40                                                                               - - Asp Ala Ser His Lys Thr Ser Ser Phe Tyr Se - #r Asn Gly Val Tyr         Tyr                                                                                              245  - #               250  - #               255              - - Gln Pro Lys Cys Gly Asn Lys Arg Gly Gln Le - #u Asp His Ala Val Leu                   260      - #           265      - #           270                   - - Val Val Gly Tyr Gly Glu Ile Asn Ser Glu Pr - #o Tyr Trp Leu Val Lys               275          - #       280          - #       285                       - - Asn Pro Gly Gln Leu Trp Gly Asn Asn Val Il - #e Phe Asp Val Ala Lys           290              - #   295              - #   300                           - - Asn Asn Asn Ala Asp Leu Thr Ile Thr Tyr Le - #u Leu Cys Thr Thr Phe       305                 3 - #10                 3 - #15                 3 -       #20                                                                               - - Lys Ile Asp Phe Tyr Gly Met                                                               325                                                             - -  - - (2) INFORMATION FOR SEQ ID NO:3:                                      - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 1109 base - #pairs                                                 (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (genomic)                                      - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:3:                                - - TTACATGCCG TAAAAATCAA TTTTAAAAGT AGTACATAGT AAATAAGTGA TC -             #GTCAAATC     60                                                                  - - CGCATTATTA TTTTTGGCGA CATCAAATAT AACATTGTTT CCCCACAATT GA -             #CCAGGATT    120                                                                  - - CTTGACCAAC CAGTAAGGTT CGCTGTTGAT TTCACCATAA CCGACTACTA AT -             #ACGGCGTG    180                                                                  - - GTCTAATTGT CCTCTTTTAT TGCCACATTT CGGTTGGTAG TAAACGCCGT TG -             #GAGTAAAA    240                                                                  - - ACTGGATGTT TTGTGTGATG CGTCAATGGC CACCGAAATC GGACCGTGTT TA -             #AGAAGGGC    300                                                                  - - AACCTTCATA GCGTCAACAT CGTTACGAGT TACGTTGACA TATCCTGTGA TG -             #GGAGCAAC    360                                                                  - - TAAAGTAACG TTGTTGACAT GGCAGTAACC ATCTTGTCCC AAGTAACCAC CA -             #TAATCCTC    420                                                                  - - TTCAGTAGGG ATTCCTCCAT GTTTCATCAT CCATTGGTAG GCGCGGAAAT CT -             #TCGCCACC    480                                                                  - - GTCGCAACCA TTATTTCCAT ATCCCCAAGA ACAATCAATC AAAGCCTGTT GA -             #GACAATCG    540                                                                  - - TACAAGATTA CCGCCGTTTT TCAAGAAATA TGCACCTTCG ATTGCTCCAA TT -             #GTTCCGAA    600                                                                  - - AGACCAGCAG GAACCACAAA CAGATTGATC TTTAACTGGA GTAACAGCTC CA -             #TAAATTCG    660                                                                  - - CCAGTCGAAA GAATCTGGTA AATTTCTTGC TTCCTTGGTG GTATTGTGAG GA -             #AAGGGTAA    720                                                                  - - ACCGCCATTA TAACCGGAGG AAGGTCTGTG TCCTCTCAAA GCCTTAAGTT CA -             #GCTTCAGT    780                                                                  - - ACGATCAGCC AAATGGTTAA CAGTCAAAGT GTATCCCAAA TTTGCACGAT TC -             #TTAGAAAA    840                                                                  - - AATGTACCTC AAGTTTAGTC TGAAAATTTC TTTGCGGAAA TGATGTTCTT CA -             #TTTTTATT    900                                                                  - - ATAAATTTTT CCGTGTTTAT TGATGAATCG GTGGAATTCT TCATCAATAT GT -             #TCGGTAAA    960                                                                  - - ATGGGGATGC ACGAATTCCC TCATTGGGGT AAAGTGCAAC TGTTCCTGAT CC -             #TGGGACCG   1020                                                                  - - GGGAAACCAA CGCAAGGTTT TTACTTCATC CGACTTCGAA AACTTCGTTA TC -             #AATTTCGC   1080                                                                  - - TTGTCTAAAA GATCGTCACC CTAATTGTA         - #                  - #               1109                                                                      - -  - - (2) INFORMATION FOR SEQ ID NO:4:                                      - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 984 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: cDNA                                               - -     (ix) FEATURE:                                                                   (A) NAME/KEY: CDS                                                              (B) LOCATION: 1..981                                                  - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:4:                                - - ATG AGG GAA TTC GTG CAT CCC CAT TTT ACC GA - #A CAT ATT GAT GAA GAA            48                                                                        Met Arg Glu Phe Val His Pro His Phe Thr Gl - #u His Ile Asp Glu Glu              1               5 - #                 10 - #                 15               - - TTC CAC CGA TTC ATC AAT AAA CAC GGA AAA AT - #T TAT AAT AAA AAT GAA            96                                                                        Phe His Arg Phe Ile Asn Lys His Gly Lys Il - #e Tyr Asn Lys Asn Glu                         20     - #             25     - #             30                   - - GAA CAT CAT TTC CGC AAA GAA ATT TTC AGA CT - #A AAC TTG AGG TAC ATT           144                                                                        Glu His His Phe Arg Lys Glu Ile Phe Arg Le - #u Asn Leu Arg Tyr Ile                     35         - #         40         - #         45                       - - TTT TCT AAG AAT CGT GCA AAT TTG GGA TAC AC - #T TTG ACT GTT AAC CAT           192                                                                        Phe Ser Lys Asn Arg Ala Asn Leu Gly Tyr Th - #r Leu Thr Val Asn His                 50             - #     55             - #     60                           - - TTG GCT GAT CGT ACT GAA GCT GAA CTT AAG GC - #T TTG AGA GGA CAC AGA           240                                                                        Leu Ala Asp Arg Thr Glu Ala Glu Leu Lys Al - #a Leu Arg Gly His Arg             65                 - # 70                 - # 75                 - # 80        - - CCT TCC TCC GGT TAT AAT GGC GGT TTA CCC TT - #T CCT CAC AAT ACC ACC           288                                                                        Pro Ser Ser Gly Tyr Asn Gly Gly Leu Pro Ph - #e Pro His Asn Thr Thr                             85 - #                 90 - #                 95               - - AAG GAA GCA AGA AAT TTA CCA GAT TCT TTC GA - #C TGG CGA ATT TAT GGA           336                                                                        Lys Glu Ala Arg Asn Leu Pro Asp Ser Phe As - #p Trp Arg Ile Tyr Gly                        100      - #           105      - #           110                   - - GCT GTT ACT CCA GTT AAA GAT CAA TCT GTT TG - #T GGT TCC TGC TGG TCT           384                                                                        Ala Val Thr Pro Val Lys Asp Gln Ser Val Cy - #s Gly Ser Cys Trp Ser                    115          - #       120          - #       125                       - - TTC GGA ACA ATT GGA GCA ATC GAA GGT GCA TA - #T TTC TTG AAA AAC GGC           432                                                                        Phe Gly Thr Ile Gly Ala Ile Glu Gly Ala Ty - #r Phe Leu Lys Asn Gly                130              - #   135              - #   140                           - - GGT AAT CTT GTA CGA TTG TCT CAA CAG GCT TT - #G ATT GAT TGT TCT TGG           480                                                                        Gly Asn Leu Val Arg Leu Ser Gln Gln Ala Le - #u Ile Asp Cys Ser Trp            145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - GGA TAT GGA AAT AAT GGT TGC GAC GGT GGC GA - #A GAT TTC CGC GCC         TAC      528                                                                     Gly Tyr Gly Asn Asn Gly Cys Asp Gly Gly Gl - #u Asp Phe Arg Ala Tyr                           165  - #               170  - #               175               - - CAA TGG ATG ATG AAA CAT GGA GGA ATC CCT AC - #T GAA GAG GAT TAT GGT           576                                                                        Gln Trp Met Met Lys His Gly Gly Ile Pro Th - #r Glu Glu Asp Tyr Gly                        180      - #           185      - #           190                   - - GGT TAC TTG GGA CAA GAT GGT TAC TGC CAT GT - #C AAC AAC GTT ACT TTA           624                                                                        Gly Tyr Leu Gly Gln Asp Gly Tyr Cys His Va - #l Asn Asn Val Thr Leu                    195          - #       200          - #       205                       - - GTT GCT CCC ATC ACA GGA TAT GTC AAC GTA AC - #T CGT AAC GAT GTT GAC           672                                                                        Val Ala Pro Ile Thr Gly Tyr Val Asn Val Th - #r Arg Asn Asp Val Asp                210              - #   215              - #   220                           - - GCT ATG AAG GTT GCC CTT CTT AAA CAC GGT CC - #G ATT TCG GTG GCC ATT           720                                                                        Ala Met Lys Val Ala Leu Leu Lys His Gly Pr - #o Ile Ser Val Ala Ile            225                 2 - #30                 2 - #35                 2 -       #40                                                                               - - GAC GCA TCA CAC AAA ACA TCC AGT TTT TAC TC - #C AAC GGC GTT TAC         TAC      768                                                                     Asp Ala Ser His Lys Thr Ser Ser Phe Tyr Se - #r Asn Gly Val Tyr Tyr                           245  - #               250  - #               255               - - CAA CCG AAA TGT GGC AAT AAA AGA GGA CAA TT - #A GAC CAC GCC GTA TTA           816                                                                        Gln Pro Lys Cys Gly Asn Lys Arg Gly Gln Le - #u Asp His Ala Val Leu                        260      - #           265      - #           270                   - - GTA GTC GGT TAT GGT GAA ATC AAC AGC GAA CC - #T TAC TGG TTG GTC AAG           864                                                                        Val Val Gly Tyr Gly Glu Ile Asn Ser Glu Pr - #o Tyr Trp Leu Val Lys                    275          - #       280          - #       285                       - - AAT CCT GGT CAA TTG TGG GGA AAC AAT GTT AT - #A TTT GAT GTC GCC AAA           912                                                                        Asn Pro Gly Gln Leu Trp Gly Asn Asn Val Il - #e Phe Asp Val Ala Lys                290              - #   295              - #   300                           - - AAT AAT AAT GCG GAT TTG ACG ATC ACT TAT TT - #A CTA TGT ACT ACT TTT           960                                                                        Asn Asn Asn Ala Asp Leu Thr Ile Thr Tyr Le - #u Leu Cys Thr Thr Phe            305                 3 - #10                 3 - #15                 3 -       #20                                                                               - - AAA ATT GAT TTT TAC GGC ATG TAA     - #                  - #                    984                                                                     Lys Ile Asp Phe Tyr Gly Met                                                                    325                                                             - -  - - (2) INFORMATION FOR SEQ ID NO:5:                                      - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 327 amino - #acids                                                 (B) TYPE: amino acid                                                           (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:5:                                - - Met Arg Glu Phe Val His Pro His Phe Thr Gl - #u His Ile Asp Glu Glu         1               5 - #                 10 - #                 15               - - Phe His Arg Phe Ile Asn Lys His Gly Lys Il - #e Tyr Asn Lys Asn Glu                    20     - #             25     - #             30                   - - Glu His His Phe Arg Lys Glu Ile Phe Arg Le - #u Asn Leu Arg Tyr Ile                35         - #         40         - #         45                       - - Phe Ser Lys Asn Arg Ala Asn Leu Gly Tyr Th - #r Leu Thr Val Asn His            50             - #     55             - #     60                           - - Leu Ala Asp Arg Thr Glu Ala Glu Leu Lys Al - #a Leu Arg Gly His Arg        65                 - # 70                 - # 75                 - # 80        - - Pro Ser Ser Gly Tyr Asn Gly Gly Leu Pro Ph - #e Pro His Asn Thr Thr                        85 - #                 90 - #                 95               - - Lys Glu Ala Arg Asn Leu Pro Asp Ser Phe As - #p Trp Arg Ile Tyr Gly                   100      - #           105      - #           110                   - - Ala Val Thr Pro Val Lys Asp Gln Ser Val Cy - #s Gly Ser Cys Trp Ser               115          - #       120          - #       125                       - - Phe Gly Thr Ile Gly Ala Ile Glu Gly Ala Ty - #r Phe Leu Lys Asn Gly           130              - #   135              - #   140                           - - Gly Asn Leu Val Arg Leu Ser Gln Gln Ala Le - #u Ile Asp Cys Ser Trp       145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - Gly Tyr Gly Asn Asn Gly Cys Asp Gly Gly Gl - #u Asp Phe Arg Ala         Tyr                                                                                              165  - #               170  - #               175              - - Gln Trp Met Met Lys His Gly Gly Ile Pro Th - #r Glu Glu Asp Tyr Gly                   180      - #           185      - #           190                   - - Gly Tyr Leu Gly Gln Asp Gly Tyr Cys His Va - #l Asn Asn Val Thr Leu               195          - #       200          - #       205                       - - Val Ala Pro Ile Thr Gly Tyr Val Asn Val Th - #r Arg Asn Asp Val Asp           210              - #   215              - #   220                           - - Ala Met Lys Val Ala Leu Leu Lys His Gly Pr - #o Ile Ser Val Ala Ile       225                 2 - #30                 2 - #35                 2 -       #40                                                                               - - Asp Ala Ser His Lys Thr Ser Ser Phe Tyr Se - #r Asn Gly Val Tyr         Tyr                                                                                              245  - #               250  - #               255              - - Gln Pro Lys Cys Gly Asn Lys Arg Gly Gln Le - #u Asp His Ala Val Leu                   260      - #           265      - #           270                   - - Val Val Gly Tyr Gly Glu Ile Asn Ser Glu Pr - #o Tyr Trp Leu Val Lys               275          - #       280          - #       285                       - - Asn Pro Gly Gln Leu Trp Gly Asn Asn Val Il - #e Phe Asp Val Ala Lys           290              - #   295              - #   300                           - - Asn Asn Asn Ala Asp Leu Thr Ile Thr Tyr Le - #u Leu Cys Thr Thr Phe       305                 3 - #10                 3 - #15                 3 -       #20                                                                               - - Lys Ile Asp Phe Tyr Gly Met                                                               325                                                             - -  - - (2) INFORMATION FOR SEQ ID NO:6:                                      - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 981 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (genomic)                                      - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:6:                                - - CATGCCGTAA AAATCAATTT TAAAAGTAGT ACATAGTAAA TAAGTGATCG TC -             #AAATCCGC     60                                                                  - - ATTATTATTT TTGGCGACAT CAAATATAAC ATTGTTTCCC CACAATTGAC CA -             #GGATTCTT    120                                                                  - - GACCAACCAG TAAGGTTCGC TGTTGATTTC ACCATAACCG ACTACTAATA CG -             #GCGTGGTC    180                                                                  - - TAATTGTCCT CTTTTATTGC CACATTTCGG TTGGTAGTAA ACGCCGTTGG AG -             #TAAAAACT    240                                                                  - - GGATGTTTTG TGTGATGCGT CAATGGCCAC CGAAATCGGA CCGTGTTTAA GA -             #AGGGCAAC    300                                                                  - - CTTCATAGCG TCAACATCGT TACGAGTTAC GTTGACATAT CCTGTGATGG GA -             #GCAACTAA    360                                                                  - - AGTAACGTTG TTGACATGGC AGTAACCATC TTGTCCCAAG TAACCACCAT AA -             #TCCTCTTC    420                                                                  - - AGTAGGGATT CCTCCATGTT TCATCATCCA TTGGTAGGCG CGGAAATCTT CG -             #CCACCGTC    480                                                                  - - GCAACCATTA TTTCCATATC CCCAAGAACA ATCAATCAAA GCCTGTTGAG AC -             #AATCGTAC    540                                                                  - - AAGATTACCG CCGTTTTTCA AGAAATATGC ACCTTCGATT GCTCCAATTG TT -             #CCGAAAGA    600                                                                  - - CCAGCAGGAA CCACAAACAG ATTGATCTTT AACTGGAGTA ACAGCTCCAT AA -             #ATTCGCCA    660                                                                  - - GTCGAAAGAA TCTGGTAAAT TTCTTGCTTC CTTGGTGGTA TTGTGAGGAA AG -             #GGTAAACC    720                                                                  - - GCCATTATAA CCGGAGGAAG GTCTGTGTCC TCTCAAAGCC TTAAGTTCAG CT -             #TCAGTACG    780                                                                  - - ATCAGCCAAA TGGTTAACAG TCAAAGTGTA TCCCAAATTT GCACGATTCT TA -             #GAAAAAAT    840                                                                  - - GTACCTCAAG TTTAGTCTGA AAATTTCTTT GCGGAAATGA TGTTCTTCAT TT -             #TTATTATA    900                                                                  - - AATTTTTCCG TGTTTATTGA TGAATCGGTG GAATTCTTCA TCAATATGTT CG -             #GTAAAATG    960                                                                  - - GGGATGCACG AATTCCCTCA T           - #                  - #                      981                                                                      - -  - - (2) INFORMATION FOR SEQ ID NO:7:                                      - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 681 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (genomic)                                      - -     (ix) FEATURE:                                                                   (A) NAME/KEY: CDS                                                              (B) LOCATION: 1..678                                                  - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:7:                                - - TTA CCA GAT TCT TTC GAC TGG CGA ATT TAT GG - #A GCT GTT ACT CCA GTT            48                                                                        Leu Pro Asp Ser Phe Asp Trp Arg Ile Tyr Gl - #y Ala Val Thr Pro Val              1               5 - #                 10 - #                 15               - - AAA GAT CAA TCT GTT TGT GGT TCC TGC TGG TC - #T TTC GGA ACA ATT GGA            96                                                                        Lys Asp Gln Ser Val Cys Gly Ser Cys Trp Se - #r Phe Gly Thr Ile Gly                         20     - #             25     - #             30                   - - GCA ATC GAA GGT GCA TAT TTC TTG AAA AAC GG - #C GGT AAT CTT GTA CGA           144                                                                        Ala Ile Glu Gly Ala Tyr Phe Leu Lys Asn Gl - #y Gly Asn Leu Val Arg                     35         - #         40         - #         45                       - - TTG TCT CAA CAG GCT TTG ATT GAT TGT TCT TG - #G GGA TAT GGA AAT AAT           192                                                                        Leu Ser Gln Gln Ala Leu Ile Asp Cys Ser Tr - #p Gly Tyr Gly Asn Asn                 50             - #     55             - #     60                           - - GGT TGC GAC GGT GGC GAA GAT TTC CGC GCC TA - #C CAA TGG ATG ATG AAA           240                                                                        Gly Cys Asp Gly Gly Glu Asp Phe Arg Ala Ty - #r Gln Trp Met Met Lys             65                 - # 70                 - # 75                 - # 80        - - CAT GGA GGA ATC CCT ACT GAA GAG GAT TAT GG - #T GGT TAC TTG GGA CAA           288                                                                        His Gly Gly Ile Pro Thr Glu Glu Asp Tyr Gl - #y Gly Tyr Leu Gly Gln                             85 - #                 90 - #                 95               - - GAT GGT TAC TGC CAT GTC AAC AAC GTT ACT TT - #A GTT GCT CCC ATC ACA           336                                                                        Asp Gly Tyr Cys His Val Asn Asn Val Thr Le - #u Val Ala Pro Ile Thr                        100      - #           105      - #           110                   - - GGA TAT GTC AAC GTA ACT CGT AAC GAT GTT GA - #C GCT ATG AAG GTT GCC           384                                                                        Gly Tyr Val Asn Val Thr Arg Asn Asp Val As - #p Ala Met Lys Val Ala                    115          - #       120          - #       125                       - - CTT CTT AAA CAC GGT CCG ATT TCG GTG GCC AT - #T GAC GCA TCA CAC AAA           432                                                                        Leu Leu Lys His Gly Pro Ile Ser Val Ala Il - #e Asp Ala Ser His Lys                130              - #   135              - #   140                           - - ACA TCC AGT TTT TAC TCC AAC GGC GTT TAC TA - #C CAA CCG AAA TGT GGC           480                                                                        Thr Ser Ser Phe Tyr Ser Asn Gly Val Tyr Ty - #r Gln Pro Lys Cys Gly            145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - AAT AAA AGA GGA CAA TTA GAC CAC GCC GTA TT - #A GTA GTC GGT TAT         GGT      528                                                                     Asn Lys Arg Gly Gln Leu Asp His Ala Val Le - #u Val Val Gly Tyr Gly                           165  - #               170  - #               175               - - GAA ATC AAC AGC GAA CCT TAC TGG TTG GTC AA - #G AAT CCT GGT CAA TTG           576                                                                        Glu Ile Asn Ser Glu Pro Tyr Trp Leu Val Ly - #s Asn Pro Gly Gln Leu                        180      - #           185      - #           190                   - - TGG GGA AAC AAT GTT ATA TTT GAT GTC GCC AA - #A AAT AAT AAT GCG GAT           624                                                                        Trp Gly Asn Asn Val Ile Phe Asp Val Ala Ly - #s Asn Asn Asn Ala Asp                    195          - #       200          - #       205                       - - TTG ACG ATC ACT TAT TTA CTA TGT ACT ACT TT - #T AAA ATT GAT TTT TAC           672                                                                        Leu Thr Ile Thr Tyr Leu Leu Cys Thr Thr Ph - #e Lys Ile Asp Phe Tyr                210              - #   215              - #   220                           - - GGC ATG TAA              - #                  - #                        - #        681                                                                   Gly Met                                                                        225                                                                             - -  - - (2) INFORMATION FOR SEQ ID NO:8:                                      - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 226 amino - #acids                                                 (B) TYPE: amino acid                                                           (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:8:                                - - Leu Pro Asp Ser Phe Asp Trp Arg Ile Tyr Gl - #y Ala Val Thr Pro         Val                                                                                1               5 - #                 10 - #                 15              - - Lys Asp Gln Ser Val Cys Gly Ser Cys Trp Se - #r Phe Gly Thr Ile Gly                    20     - #             25     - #             30                   - - Ala Ile Glu Gly Ala Tyr Phe Leu Lys Asn Gl - #y Gly Asn Leu Val Arg                35         - #         40         - #         45                       - - Leu Ser Gln Gln Ala Leu Ile Asp Cys Ser Tr - #p Gly Tyr Gly Asn Asn            50             - #     55             - #     60                           - - Gly Cys Asp Gly Gly Glu Asp Phe Arg Ala Ty - #r Gln Trp Met Met Lys        65                 - # 70                 - # 75                 - # 80        - - His Gly Gly Ile Pro Thr Glu Glu Asp Tyr Gl - #y Gly Tyr Leu Gly Gln                        85 - #                 90 - #                 95               - - Asp Gly Tyr Cys His Val Asn Asn Val Thr Le - #u Val Ala Pro Ile Thr                   100      - #           105      - #           110                   - - Gly Tyr Val Asn Val Thr Arg Asn Asp Val As - #p Ala Met Lys Val Ala               115          - #       120          - #       125                       - - Leu Leu Lys His Gly Pro Ile Ser Val Ala Il - #e Asp Ala Ser His Lys           130              - #   135              - #   140                           - - Thr Ser Ser Phe Tyr Ser Asn Gly Val Tyr Ty - #r Gln Pro Lys Cys Gly       145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - Asn Lys Arg Gly Gln Leu Asp His Ala Val Le - #u Val Val Gly Tyr         Gly                                                                                              165  - #               170  - #               175              - - Glu Ile Asn Ser Glu Pro Tyr Trp Leu Val Ly - #s Asn Pro Gly Gln Leu                   180      - #           185      - #           190                   - - Trp Gly Asn Asn Val Ile Phe Asp Val Ala Ly - #s Asn Asn Asn Ala Asp               195          - #       200          - #       205                       - - Leu Thr Ile Thr Tyr Leu Leu Cys Thr Thr Ph - #e Lys Ile Asp Phe Tyr           210              - #   215              - #   220                           - - Gly Met                                                                   225                                                                             - -  - - (2) INFORMATION FOR SEQ ID NO:9:                                      - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 775 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: cDNA                                               - -     (ix) FEATURE:                                                                   (A) NAME/KEY: CDS                                                              (B) LOCATION: 1..685                                                  - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #any amino acid                                          (B) LOCATION: 187                                                     - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:9:                                - - TCA AAT CGG ATT GTT AAT GGA GTT AAT GCC AA - #A AAC GGT TCT GCT CCA            48                                                                        Ser Asn Arg Ile Val Asn Gly Val Asn Ala Ly - #s Asn Gly Ser Ala Pro              1               5 - #                 10 - #                 15               - - TAT ATG GCT TCT CTA AGA GAT GTT ATG GAA AC - #C ATT TCT GTG GAG CAT            96                                                                        Tyr Met Ala Ser Leu Arg Asp Val Met Glu Th - #r Ile Ser Val Glu His                         20     - #             25     - #             30                   - - CGA TAT TGG ATG AAC CGC TGG ATT CTT ACT GC - #T GCC CAT TGC CTT ACT           144                                                                        Arg Tyr Trp Met Asn Arg Trp Ile Leu Thr Al - #a Ala His Cys Leu Thr                     35         - #         40         - #         45                       - - GAC GGT TAT CTA GAT ACA GTC TAC GTT GGT TC - #A AAT CAT CTT TCT GGC           192                                                                        Asp Gly Tyr Leu Asp Thr Val Tyr Val Gly Se - #r Asn His Leu Ser Gly                 50             - #     55             - #     60                           - - GAC GGA GAG TAC TAC AAT GTA GAA GAA CAA GT - #C ATC CAT GAT AAA TAT           240                                                                        Asp Gly Glu Tyr Tyr Asn Val Glu Glu Gln Va - #l Ile His Asp Lys Tyr             65                 - # 70                 - # 75                 - # 80        - - TTT GGT CAA ACA ACC GGC TTC AAA AAT GAT AT - #T GCT CTC GTC AAA GTT           288                                                                        Phe Gly Gln Thr Thr Gly Phe Lys Asn Asp Il - #e Ala Leu Val Lys Val                             85 - #                 90 - #                 95               - - TCT AGT GCT ATA AAA CTT AGC AAA AAT GTT CG - #T CCC ATC AAA TTG CAC           336                                                                        Ser Ser Ala Ile Lys Leu Ser Lys Asn Val Ar - #g Pro Ile Lys Leu His                        100      - #           105      - #           110                   - - AAA GAT TTT ATA CGC GGA GGT GAA AAA TTG AA - #A ATT ACT GGA TGG GGA           384                                                                        Lys Asp Phe Ile Arg Gly Gly Glu Lys Leu Ly - #s Ile Thr Gly Trp Gly                    115          - #       120          - #       125                       - - TTG ACC AAT CAA ACT CAT GGT GAA GTT CCT GA - #T GCT CTT CAA GAG TTA           432                                                                        Leu Thr Asn Gln Thr His Gly Glu Val Pro As - #p Ala Leu Gln Glu Leu                130              - #   135              - #   140                           - - CAG GTA GAA GCA CTT TCT AAC TCT AAA TGC AA - #G GCA ATT ACT GGT GTC           480                                                                        Gln Val Glu Ala Leu Ser Asn Ser Lys Cys Ly - #s Ala Ile Thr Gly Val            145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - CAT CTT CCT GCT CAT CTC TGC ACC TTC AAA GC - #A CCT CAA AAG GGT         GTA      528                                                                     His Leu Pro Ala His Leu Cys Thr Phe Lys Al - #a Pro Gln Lys Gly Val                           165  - #               170  - #               175               - - TGC ATG GGT GAC TCT GGT GGT CCT CTG GTC NA - #T AAG GGC AAG CAA GTT           576                                                                        Cys Met Gly Asp Ser Gly Gly Pro Leu Val Xa - #a Lys Gly Lys Gln Val                        180      - #           185      - #           190                   - - GGA GTC ACA TCT TTC GTC TGG GAA GGT TGT GC - #T TTG GGC AAC CCT GAT           624                                                                        Gly Val Thr Ser Phe Val Trp Glu Gly Cys Al - #a Leu Gly Asn Pro Asp                    195          - #       200          - #       205                       - - TTC TTT ACA AGA GTT TCG CTT TAT GTA GAC TG - #G GTC AAA AAG ATT CAA           672                                                                        Phe Phe Thr Arg Val Ser Leu Tyr Val Asp Tr - #p Val Lys Lys Ile Gln                210              - #   215              - #   220                           - - AAA GAA TAT AAA T GATATGTTGA TTGTCACTAA AATGCATCGA - #TTTGGATAAT             725                                                                         Lys Glu Tyr Lys                                                                225                                                                             - - TTGGTTGTGA ATATAATTTT ATTTCTAGCA TCAAAAAAAA AAAAAAAAAA  - #                  775                                                                          - -  - - (2) INFORMATION FOR SEQ ID NO:10:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 228 amino - #acids                                                 (B) TYPE: amino acid                                                           (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #any amino acid                                          (B) LOCATION: 187                                                     - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:10:                               - - Ser Asn Arg Ile Val Asn Gly Val Asn Ala Ly - #s Asn Gly Ser Ala Pro         1               5 - #                 10 - #                 15               - - Tyr Met Ala Ser Leu Arg Asp Val Met Glu Th - #r Ile Ser Val Glu His                    20     - #             25     - #             30                   - - Arg Tyr Trp Met Asn Arg Trp Ile Leu Thr Al - #a Ala His Cys Leu Thr                35         - #         40         - #         45                       - - Asp Gly Tyr Leu Asp Thr Val Tyr Val Gly Se - #r Asn His Leu Ser Gly            50             - #     55             - #     60                           - - Asp Gly Glu Tyr Tyr Asn Val Glu Glu Gln Va - #l Ile His Asp Lys Tyr        65                 - # 70                 - # 75                 - # 80        - - Phe Gly Gln Thr Thr Gly Phe Lys Asn Asp Il - #e Ala Leu Val Lys Val                        85 - #                 90 - #                 95               - - Ser Ser Ala Ile Lys Leu Ser Lys Asn Val Ar - #g Pro Ile Lys Leu His                   100      - #           105      - #           110                   - - Lys Asp Phe Ile Arg Gly Gly Glu Lys Leu Ly - #s Ile Thr Gly Trp Gly               115          - #       120          - #       125                       - - Leu Thr Asn Gln Thr His Gly Glu Val Pro As - #p Ala Leu Gln Glu Leu           130              - #   135              - #   140                           - - Gln Val Glu Ala Leu Ser Asn Ser Lys Cys Ly - #s Ala Ile Thr Gly Val       145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - His Leu Pro Ala His Leu Cys Thr Phe Lys Al - #a Pro Gln Lys Gly         Val                                                                                              165  - #               170  - #               175              - - Cys Met Gly Asp Ser Gly Gly Pro Leu Val Xa - #a Lys Gly Lys Gln Val                   180      - #           185      - #           190                   - - Gly Val Thr Ser Phe Val Trp Glu Gly Cys Al - #a Leu Gly Asn Pro Asp               195          - #       200          - #       205                       - - Phe Phe Thr Arg Val Ser Leu Tyr Val Asp Tr - #p Val Lys Lys Ile Gln           210              - #   215              - #   220                           - - Lys Glu Tyr Lys                                                           225                                                                             - -  - - (2) INFORMATION FOR SEQ ID NO:11:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 775 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (genomic)                                      - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:11:                               - - TTTTTTTTTT TTTTTTTTGA TGCTAGAAAT AAAATTATAT TCACAACCAA AT -              #TATCCAAA     60                                                                  - - TCGATGCATT TTAGTGACAA TCAACATATC ATTTATATTC TTTTTGAATC TT -             #TTTGACCC    120                                                                  - - AGTCTACATA AAGCGAAACT CTTGTAAAGA AATCAGGGTT GCCCAAAGCA CA -             #ACCTTCCC    180                                                                  - - AGACGAAAGA TGTGACTCCA ACTTGCTTGC CCTTATNGAC CAGAGGACCA CC -             #AGAGTCAC    240                                                                  - - CCATGCATAC ACCCTTTTGA GGTGCTTTGA AGGTGCAGAG ATGAGCAGGA AG -             #ATGGACAC    300                                                                  - - CAGTAATTGC CTTGCATTTA GAGTTAGAAA GTGCTTCTAC CTGTAACTCT TG -             #AAGAGCAT    360                                                                  - - CAGGAACTTC ACCATGAGTT TGATTGGTCA ATCCCCATCC AGTAATTTTC AA -             #TTTTTCAC    420                                                                  - - CTCCGCGTAT AAAATCTTTG TGCAATTTGA TGGGACGAAC ATTTTTGCTA AG -             #TTTTATAG    480                                                                  - - CACTAGAAAC TTTGACGAGA GCAATATCAT TTTTGAAGCC GGTTGTTTGA CC -             #AAAATATT    540                                                                  - - TATCATGGAT GACTTGTTCT TCTACATTGT AGTACTCTCC GTCGCCAGAA AG -             #ATGATTTG    600                                                                  - - AACCAACGTA GACTGTATCT AGATAACCGT CAGTAAGGCA ATGGGCAGCA GT -             #AAGAATCC    660                                                                  - - AGCGGTTCAT CCAATATCGA TGCTCCACAG AAATGGTTTC CATAACATCT CT -             #TAGAGAAG    720                                                                  - - CCATATATGG AGCAGAACCG TTTTTGGCAT TAACTCCATT AACAATCCGA TT - #TGA              775                                                                        - -  - - (2) INFORMATION FOR SEQ ID NO:12:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 675 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (genomic)                                      - -     (ix) FEATURE:                                                                   (A) NAME/KEY: CDS                                                              (B) LOCATION: 1..675                                                  - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #any amino acid                                          (B) LOCATION: 184                                                     - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:12:                               - - ATT GTT AAT GGA GTT AAT GCC AAA AAC GGT TC - #T GCT CCA TAT ATG GCT            48                                                                        Ile Val Asn Gly Val Asn Ala Lys Asn Gly Se - #r Ala Pro Tyr Met Ala              1               5 - #                 10 - #                 15               - - TCT CTA AGA GAT GTT ATG GAA ACC ATT TCT GT - #G GAG CAT CGA TAT TGG            96                                                                        Ser Leu Arg Asp Val Met Glu Thr Ile Ser Va - #l Glu His Arg Tyr Trp                         20     - #             25     - #             30                   - - ATG AAC CGC TGG ATT CTT ACT GCT GCC CAT TG - #C CTT ACT GAC GGT TAT           144                                                                        Met Asn Arg Trp Ile Leu Thr Ala Ala His Cy - #s Leu Thr Asp Gly Tyr                     35         - #         40         - #         45                       - - CTA GAT ACA GTC TAC GTT GGT TCA AAT CAT CT - #T TCT GGC GAC GGA GAG           192                                                                        Leu Asp Thr Val Tyr Val Gly Ser Asn His Le - #u Ser Gly Asp Gly Glu                 50             - #     55             - #     60                           - - TAC TAC AAT GTA GAA GAA CAA GTC ATC CAT GA - #T AAA TAT TTT GGT CAA           240                                                                        Tyr Tyr Asn Val Glu Glu Gln Val Ile His As - #p Lys Tyr Phe Gly Gln             65                 - # 70                 - # 75                 - # 80        - - ACA ACC GGC TTC AAA AAT GAT ATT GCT CTC GT - #C AAA GTT TCT AGT GCT           288                                                                        Thr Thr Gly Phe Lys Asn Asp Ile Ala Leu Va - #l Lys Val Ser Ser Ala                             85 - #                 90 - #                 95               - - ATA AAA CTT AGC AAA AAT GTT CGT CCC ATC AA - #A TTG CAC AAA GAT TTT           336                                                                        Ile Lys Leu Ser Lys Asn Val Arg Pro Ile Ly - #s Leu His Lys Asp Phe                        100      - #           105      - #           110                   - - ATA CGC GGA GGT GAA AAA TTG AAA ATT ACT GG - #A TGG GGA TTG ACC AAT           384                                                                        Ile Arg Gly Gly Glu Lys Leu Lys Ile Thr Gl - #y Trp Gly Leu Thr Asn                    115          - #       120          - #       125                       - - CAA ACT CAT GGT GAA GTT CCT GAT GCT CTT CA - #A GAG TTA CAG GTA GAA           432                                                                        Gln Thr His Gly Glu Val Pro Asp Ala Leu Gl - #n Glu Leu Gln Val Glu                130              - #   135              - #   140                           - - GCA CTT TCT AAC TCT AAA TGC AAG GCA ATT AC - #T GGT GTC CAT CTT CCT           480                                                                        Ala Leu Ser Asn Ser Lys Cys Lys Ala Ile Th - #r Gly Val His Leu Pro            145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - GCT CAT CTC TGC ACC TTC AAA GCA CCT CAA AA - #G GGT GTA TGC ATG         GGT      528                                                                     Ala His Leu Cys Thr Phe Lys Ala Pro Gln Ly - #s Gly Val Cys Met Gly                           165  - #               170  - #               175               - - GAC TCT GGT GGT CCT CTG GTC NAT AAG GGC AA - #G CAA GTT GGA GTC ACA           576                                                                        Asp Ser Gly Gly Pro Leu Val Xaa Lys Gly Ly - #s Gln Val Gly Val Thr                        180      - #           185      - #           190                   - - TCT TTC GTC TGG GAA GGT TGT GCT TTG GGC AA - #C CCT GAT TTC TTT ACA           624                                                                        Ser Phe Val Trp Glu Gly Cys Ala Leu Gly As - #n Pro Asp Phe Phe Thr                    195          - #       200          - #       205                       - - AGA GTT TCG CTT TAT GTA GAC TGG GTC AAA AA - #G ATT CAA AAA GAA TAT           672                                                                        Arg Val Ser Leu Tyr Val Asp Trp Val Lys Ly - #s Ile Gln Lys Glu Tyr                210              - #   215              - #   220                           - - AAA                  - #                  - #                  - #                 675                                                                   Lys                                                                            225                                                                             - -  - - (2) INFORMATION FOR SEQ ID NO:13:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 225 amino - #acids                                                 (B) TYPE: amino acid                                                           (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #any amino acid                                          (B) LOCATION: 184                                                     - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:13:                               - - Ile Val Asn Gly Val Asn Ala Lys Asn Gly Se - #r Ala Pro Tyr Met Ala         1               5 - #                 10 - #                 15               - - Ser Leu Arg Asp Val Met Glu Thr Ile Ser Va - #l Glu His Arg Tyr Trp                    20     - #             25     - #             30                   - - Met Asn Arg Trp Ile Leu Thr Ala Ala His Cy - #s Leu Thr Asp Gly Tyr                35         - #         40         - #         45                       - - Leu Asp Thr Val Tyr Val Gly Ser Asn His Le - #u Ser Gly Asp Gly Glu            50             - #     55             - #     60                           - - Tyr Tyr Asn Val Glu Glu Gln Val Ile His As - #p Lys Tyr Phe Gly Gln        65                 - # 70                 - # 75                 - # 80        - - Thr Thr Gly Phe Lys Asn Asp Ile Ala Leu Va - #l Lys Val Ser Ser Ala                        85 - #                 90 - #                 95               - - Ile Lys Leu Ser Lys Asn Val Arg Pro Ile Ly - #s Leu His Lys Asp Phe                   100      - #           105      - #           110                   - - Ile Arg Gly Gly Glu Lys Leu Lys Ile Thr Gl - #y Trp Gly Leu Thr Asn               115          - #       120          - #       125                       - - Gln Thr His Gly Glu Val Pro Asp Ala Leu Gl - #n Glu Leu Gln Val Glu           130              - #   135              - #   140                           - - Ala Leu Ser Asn Ser Lys Cys Lys Ala Ile Th - #r Gly Val His Leu Pro       145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - Ala His Leu Cys Thr Phe Lys Ala Pro Gln Ly - #s Gly Val Cys Met         Gly                                                                                              165  - #               170  - #               175              - - Asp Ser Gly Gly Pro Leu Val Xaa Lys Gly Ly - #s Gln Val Gly Val Thr                   180      - #           185      - #           190                   - - Ser Phe Val Trp Glu Gly Cys Ala Leu Gly As - #n Pro Asp Phe Phe Thr               195          - #       200          - #       205                       - - Arg Val Ser Leu Tyr Val Asp Trp Val Lys Ly - #s Ile Gln Lys Glu Tyr           210              - #   215              - #   220                           - - Lys                                                                       225                                                                             - -  - - (2) INFORMATION FOR SEQ ID NO:14:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 675 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (genomic)                                      - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:14:                               - - TTTATATTCT TTTTGAATCT TTTTGACCCA GTCTACATAA AGCGAAACTC TT -              #GTAAAGAA     60                                                                  - - ATCAGGGTTG CCCAAAGCAC AACCTTCCCA GACGAAAGAT GTGACTCCAA CT -             #TGCTTGCC    120                                                                  - - CTTATNGACC AGAGGACCAC CAGAGTCACC CATGCATACA CCCTTTTGAG GT -             #GCTTTGAA    180                                                                  - - GGTGCAGAGA TGAGCAGGAA GATGGACACC AGTAATTGCC TTGCATTTAG AG -             #TTAGAAAG    240                                                                  - - TGCTTCTACC TGTAACTCTT GAAGAGCATC AGGAACTTCA CCATGAGTTT GA -             #TTGGTCAA    300                                                                  - - TCCCCATCCA GTAATTTTCA ATTTTTCACC TCCGCGTATA AAATCTTTGT GC -             #AATTTGAT    360                                                                  - - GGGACGAACA TTTTTGCTAA GTTTTATAGC ACTAGAAACT TTGACGAGAG CA -             #ATATCATT    420                                                                  - - TTTGAAGCCG GTTGTTTGAC CAAAATATTT ATCATGGATG ACTTGTTCTT CT -             #ACATTGTA    480                                                                  - - GTACTCTCCG TCGCCAGAAA GATGATTTGA ACCAACGTAG ACTGTATCTA GA -             #TAACCGTC    540                                                                  - - AGTAAGGCAA TGGGCAGCAG TAAGAATCCA GCGGTTCATC CAATATCGAT GC -             #TCCACAGA    600                                                                  - - AATGGTTTCC ATAACATCTC TTAGAGAAGC CATATATGGA GCAGAACCGT TT -             #TTGGCATT    660                                                                  - - AACTCCATTA ACAAT              - #                  - #                       - #   675                                                                   - -  - - (2) INFORMATION FOR SEQ ID NO:15:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 1089 base - #pairs                                                 (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: cDNA                                               - -     (ix) FEATURE:                                                                   (A) NAME/KEY: CDS                                                              (B) LOCATION: 33..807                                                 - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:15:                               - - GTTTTTAGTG CGGTGTTAAA TAAAAGTTTA AA ATG AAA CTC ATC - # ATC GTG CTA            53                                                                                          - #                  - #Met Lys Leu Ile Ile Val Leu                            - #                  - #  1               5                   - - GCA TTT GTT TTA GGA ATT TGT TCA GGT TCT CC - #A CAT TCA AGA ATA ATT           101                                                                        Ala Phe Val Leu Gly Ile Cys Ser Gly Ser Pr - #o His Ser Arg Ile Ile                     10         - #         15         - #         20                       - - TGT GGT CAA AAT GCC AAA AAA AAT TCG GCT CC - #A TAC ATG GCA TCG GTT           149                                                                        Cys Gly Gln Asn Ala Lys Lys Asn Ser Ala Pr - #o Tyr Met Ala Ser Val                 25             - #     30             - #     35                           - - CAA CTT TTA GAT AAA GTT GAA GGA GTC GAA AA - #A TTG TTT CAT TTT TGC           197                                                                        Gln Leu Leu Asp Lys Val Glu Gly Val Glu Ly - #s Leu Phe His Phe Cys             40                 - # 45                 - # 50                 - # 55        - - GGA GGA GCA ATA GTT AAT GAT AGA TGG ATT TT - #G ACT GCT GCA CAT TGT           245                                                                        Gly Gly Ala Ile Val Asn Asp Arg Trp Ile Le - #u Thr Ala Ala His Cys                             60 - #                 65 - #                 70               - - TTG AGA GGC AAA GAC CAC CTC CTG GAC AAA CT - #G TTC ATT GCA GTC GGC           293                                                                        Leu Arg Gly Lys Asp His Leu Leu Asp Lys Le - #u Phe Ile Ala Val Gly                         75     - #             80     - #             85                   - - CTG ACA AAT TTA GGT GAA GGA GGC ACC GTG TA - #T CCT GTA GAA AAA GGC           341                                                                        Leu Thr Asn Leu Gly Glu Gly Gly Thr Val Ty - #r Pro Val Glu Lys Gly                     90         - #         95         - #        100                       - - ATC ATG CAC GAA GAA TAT GAA CAT TAT GAC AT - #A GTC AAC GAT ATT GCA           389                                                                        Ile Met His Glu Glu Tyr Glu His Tyr Asp Il - #e Val Asn Asp Ile Ala                105              - #   110              - #   115                           - - CTA ATC AAA GTC AAA TCT CCG ATA GAA TTC AA - #T GAA AAA GTA ACG ACT           437                                                                        Leu Ile Lys Val Lys Ser Pro Ile Glu Phe As - #n Glu Lys Val Thr Thr            120                 1 - #25                 1 - #30                 1 -       #35                                                                               - - GTA AAA TTA GGT GAG GAT TAT GTT GGC GGA GA - #C GTC CAA CTT CGA         TTG      485                                                                     Val Lys Leu Gly Glu Asp Tyr Val Gly Gly As - #p Val Gln Leu Arg Leu                           140  - #               145  - #               150               - - ACA GGA TGG GGA GTT ACG ACA AAT GAG GGA AT - #C GGA AGC CCG AGT CAA           533                                                                        Thr Gly Trp Gly Val Thr Thr Asn Glu Gly Il - #e Gly Ser Pro Ser Gln                        155      - #           160      - #           165                   - - AAA TTA CAG GTC ATG ACA GCC AAA TCA CTA AC - #T TAT GAG GAT TGC AAA           581                                                                        Lys Leu Gln Val Met Thr Ala Lys Ser Leu Th - #r Tyr Glu Asp Cys Lys                    170          - #       175          - #       180                       - - AAC GCA ATT TAT AAA AAG ACT TTC GAA AGC CA - #A ATT TGT GCA CAG GCT           629                                                                        Asn Ala Ile Tyr Lys Lys Thr Phe Glu Ser Gl - #n Ile Cys Ala Gln Ala                185              - #   190              - #   195                           - - AAA AAA GGA ACC GGA TCT TGT AAG GGT GAT TC - #T GGT GGT CCA TTA GTC           677                                                                        Lys Lys Gly Thr Gly Ser Cys Lys Gly Asp Se - #r Gly Gly Pro Leu Val            200                 2 - #05                 2 - #10                 2 -       #15                                                                               - - CAA GGA AAC AAT ACA TTG GTC GGT TTA GTA TC - #C TGG GGT ATG CAA         CCT      725                                                                     Gln Gly Asn Asn Thr Leu Val Gly Leu Val Se - #r Trp Gly Met Gln Pro                           220  - #               225  - #               230               - - TGT GGA AGT GGT TAT TAT CCT GAC GTT TAC AC - #A AGA ATT ACA TCG TTT           773                                                                        Cys Gly Ser Gly Tyr Tyr Pro Asp Val Tyr Th - #r Arg Ile Thr Ser Phe                        235      - #           240      - #           245                   - - TTG GAC TGG ATT AAC ACG ACA ATG TCA GAA AA - #T T AAAGANAAAA                  - #817                                                                     Leu Asp Trp Ile Asn Thr Thr Met Ser Glu As - #n                                        250          - #       255                                              - - AANTATCAAA ATAGTAGTTA AAATATTTTG TNGACTACTG TAAAAGTATT GA -              #AATTAACA    877                                                                  - - AATATTTGTT TTGTATATAG GGCTGGTACC TAAAGACATT ATTATTTGTT AA -             #AACTGATA    937                                                                  - - TTTATTTTTA TGACGAATTT CTATATTTAT AATATTATTT TATATGTATG TA -             #TTTGATTG    997                                                                  - - TATTTTAAAT ATGGACTGGT TGTTCCCAAT GTTATAAATT AAAATTATAA AA -             #ATACATCT   1057                                                                  - - ACTATTTTAT ACTAAAAAAA AAAAAAAAAA AA       - #                  - #             1089                                                                      - -  - - (2) INFORMATION FOR SEQ ID NO:16:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 258 amino - #acids                                                 (B) TYPE: amino acid                                                           (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:16:                               - - Met Lys Leu Ile Ile Val Leu Ala Phe Val Le - #u Gly Ile Cys Ser Gly         1               5 - #                 10 - #                 15               - - Ser Pro His Ser Arg Ile Ile Cys Gly Gln As - #n Ala Lys Lys Asn Ser                    20     - #             25     - #             30                   - - Ala Pro Tyr Met Ala Ser Val Gln Leu Leu As - #p Lys Val Glu Gly Val                35         - #         40         - #         45                       - - Glu Lys Leu Phe His Phe Cys Gly Gly Ala Il - #e Val Asn Asp Arg Trp            50             - #     55             - #     60                           - - Ile Leu Thr Ala Ala His Cys Leu Arg Gly Ly - #s Asp His Leu Leu Asp        65                 - # 70                 - # 75                 - # 80        - - Lys Leu Phe Ile Ala Val Gly Leu Thr Asn Le - #u Gly Glu Gly Gly Thr                        85 - #                 90 - #                 95               - - Val Tyr Pro Val Glu Lys Gly Ile Met His Gl - #u Glu Tyr Glu His Tyr                   100      - #           105      - #           110                   - - Asp Ile Val Asn Asp Ile Ala Leu Ile Lys Va - #l Lys Ser Pro Ile Glu               115          - #       120          - #       125                       - - Phe Asn Glu Lys Val Thr Thr Val Lys Leu Gl - #y Glu Asp Tyr Val Gly           130              - #   135              - #   140                           - - Gly Asp Val Gln Leu Arg Leu Thr Gly Trp Gl - #y Val Thr Thr Asn Glu       145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - Gly Ile Gly Ser Pro Ser Gln Lys Leu Gln Va - #l Met Thr Ala Lys         Ser                                                                                              165  - #               170  - #               175              - - Leu Thr Tyr Glu Asp Cys Lys Asn Ala Ile Ty - #r Lys Lys Thr Phe Glu                   180      - #           185      - #           190                   - - Ser Gln Ile Cys Ala Gln Ala Lys Lys Gly Th - #r Gly Ser Cys Lys Gly               195          - #       200          - #       205                       - - Asp Ser Gly Gly Pro Leu Val Gln Gly Asn As - #n Thr Leu Val Gly Leu           210              - #   215              - #   220                           - - Val Ser Trp Gly Met Gln Pro Cys Gly Ser Gl - #y Tyr Tyr Pro Asp Val       225                 2 - #30                 2 - #35                 2 -       #40                                                                               - - Tyr Thr Arg Ile Thr Ser Phe Leu Asp Trp Il - #e Asn Thr Thr Met         Ser                                                                                              245  - #               250  - #               255              - - Glu Asn                                                                    - -  - - (2) INFORMATION FOR SEQ ID NO:17:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 1089 base - #pairs                                                 (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (genomic)                                      - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:17:                               - - TTTTTTTTTT TTTTTTTTTA GTATAAAATA GTAGATGTAT TTTTATAATT TT -              #AATTTATA     60                                                                  - - ACATTGGGAA CAACCAGTCC ATATTTAAAA TACAATCAAA TACATACATA TA -             #AAATAATA    120                                                                  - - TTATAAATAT AGAAATTCGT CATAAAAATA AATATCAGTT TTAACAAATA AT -             #AATGTCTT    180                                                                  - - TAGGTACCAG CCCTATATAC AAAACAAATA TTTGTTAATT TCAATACTTT TA -             #CAGTAGTC    240                                                                  - - NACAAAATAT TTTAACTACT ATTTTGATAN TTTTTTNTCT TTAATTTTCT GA -             #CATTGTCG    300                                                                  - - TGTTAATCCA GTCCAAAAAC GATGTAATTC TTGTGTAAAC GTCAGGATAA TA -             #ACCACTTC    360                                                                  - - CACAAGGTTG CATACCCCAG GATACTAAAC CGACCAATGT ATTGTTTCCT TG -             #GACTAATG    420                                                                  - - GACCACCAGA ATCACCCTTA CAAGATCCGG TTCCTTTTTT AGCCTGTGCA CA -             #AATTTGGC    480                                                                  - - TTTCGAAAGT CTTTTTATAA ATTGCGTTTT TGCAATCCTC ATAAGTTAGT GA -             #TTTGGCTG    540                                                                  - - TCATGACCTG TAATTTTTGA CTCGGGCTTC CGATTCCCTC ATTTGTCGTA AC -             #TCCCCATC    600                                                                  - - CTGTCAATCG AAGTTGGACG TCTCCGCCAA CATAATCCTC ACCTAATTTT AC -             #AGTCGTTA    660                                                                  - - CTTTTTCATT GAATTCTATC GGAGATTTGA CTTTGATTAG TGCAATATCG TT -             #GACTATGT    720                                                                  - - CATAATGTTC ATATTCTTCG TGCATGATGC CTTTTTCTAC AGGATACACG GT -             #GCCTCCTT    780                                                                  - - CACCTAAATT TGTCAGGCCG ACTGCAATGA ACAGTTTGTC CAGGAGGTGG TC -             #TTTGCCTC    840                                                                  - - TCAAACAATG TGCAGCAGTC AAAATCCATC TATCATTAAC TATTGCTCCT CC -             #GCAAAAAT    900                                                                  - - GAAACAATTT TTCGACTCCT TCAACTTTAT CTAAAAGTTG AACCGATGCC AT -             #GTATGGAG    960                                                                  - - CCGAATTTTT TTTGGCATTT TGACCACAAA TTATTCTTGA ATGTGGAGAA CC -             #TGAACAAA   1020                                                                  - - TTCCTAAAAC AAATGCTAGC ACGATGATGA GTTTCATTTT AAACTTTTAT TT -             #AACACCGC   1080                                                                  - - ACTAAAAAC                - #                  - #                       - #       1089                                                                   - -  - - (2) INFORMATION FOR SEQ ID NO:18:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 774 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (genomic)                                      - -     (ix) FEATURE:                                                                   (A) NAME/KEY: CDS                                                              (B) LOCATION: 1..774                                                  - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:18:                               - - ATG AAA CTC ATC ATC GTG CTA GCA TTT GTT TT - #A GGA ATT TGT TCA GGT            48                                                                        Met Lys Leu Ile Ile Val Leu Ala Phe Val Le - #u Gly Ile Cys Ser Gly              1               5 - #                 10 - #                 15               - - TCT CCA CAT TCA AGA ATA ATT TGT GGT CAA AA - #T GCC AAA AAA AAT TCG            96                                                                        Ser Pro His Ser Arg Ile Ile Cys Gly Gln As - #n Ala Lys Lys Asn Ser                         20     - #             25     - #             30                   - - GCT CCA TAC ATG GCA TCG GTT CAA CTT TTA GA - #T AAA GTT GAA GGA GTC           144                                                                        Ala Pro Tyr Met Ala Ser Val Gln Leu Leu As - #p Lys Val Glu Gly Val                     35         - #         40         - #         45                       - - GAA AAA TTG TTT CAT TTT TGC GGA GGA GCA AT - #A GTT AAT GAT AGA TGG           192                                                                        Glu Lys Leu Phe His Phe Cys Gly Gly Ala Il - #e Val Asn Asp Arg Trp                 50             - #     55             - #     60                           - - ATT TTG ACT GCT GCA CAT TGT TTG AGA GGC AA - #A GAC CAC CTC CTG GAC           240                                                                        Ile Leu Thr Ala Ala His Cys Leu Arg Gly Ly - #s Asp His Leu Leu Asp             65                 - # 70                 - # 75                 - # 80        - - AAA CTG TTC ATT GCA GTC GGC CTG ACA AAT TT - #A GGT GAA GGA GGC ACC           288                                                                        Lys Leu Phe Ile Ala Val Gly Leu Thr Asn Le - #u Gly Glu Gly Gly Thr                             85 - #                 90 - #                 95               - - GTG TAT CCT GTA GAA AAA GGC ATC ATG CAC GA - #A GAA TAT GAA CAT TAT           336                                                                        Val Tyr Pro Val Glu Lys Gly Ile Met His Gl - #u Glu Tyr Glu His Tyr                        100      - #           105      - #           110                   - - GAC ATA GTC AAC GAT ATT GCA CTA ATC AAA GT - #C AAA TCT CCG ATA GAA           384                                                                        Asp Ile Val Asn Asp Ile Ala Leu Ile Lys Va - #l Lys Ser Pro Ile Glu                    115          - #       120          - #       125                       - - TTC AAT GAA AAA GTA ACG ACT GTA AAA TTA GG - #T GAG GAT TAT GTT GGC           432                                                                        Phe Asn Glu Lys Val Thr Thr Val Lys Leu Gl - #y Glu Asp Tyr Val Gly                130              - #   135              - #   140                           - - GGA GAC GTC CAA CTT CGA TTG ACA GGA TGG GG - #A GTT ACG ACA AAT GAG           480                                                                        Gly Asp Val Gln Leu Arg Leu Thr Gly Trp Gl - #y Val Thr Thr Asn Glu            145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - GGA ATC GGA AGC CCG AGT CAA AAA TTA CAG GT - #C ATG ACA GCC AAA         TCA      528                                                                     Gly Ile Gly Ser Pro Ser Gln Lys Leu Gln Va - #l Met Thr Ala Lys Ser                           165  - #               170  - #               175               - - CTA ACT TAT GAG GAT TGC AAA AAC GCA ATT TA - #T AAA AAG ACT TTC GAA           576                                                                        Leu Thr Tyr Glu Asp Cys Lys Asn Ala Ile Ty - #r Lys Lys Thr Phe Glu                        180      - #           185      - #           190                   - - AGC CAA ATT TGT GCA CAG GCT AAA AAA GGA AC - #C GGA TCT TGT AAG GGT           624                                                                        Ser Gln Ile Cys Ala Gln Ala Lys Lys Gly Th - #r Gly Ser Cys Lys Gly                    195          - #       200          - #       205                       - - GAT TCT GGT GGT CCA TTA GTC CAA GGA AAC AA - #T ACA TTG GTC GGT TTA           672                                                                        Asp Ser Gly Gly Pro Leu Val Gln Gly Asn As - #n Thr Leu Val Gly Leu                210              - #   215              - #   220                           - - GTA TCC TGG GGT ATG CAA CCT TGT GGA AGT GG - #T TAT TAT CCT GAC GTT           720                                                                        Val Ser Trp Gly Met Gln Pro Cys Gly Ser Gl - #y Tyr Tyr Pro Asp Val            225                 2 - #30                 2 - #35                 2 -       #40                                                                               - - TAC ACA AGA ATT ACA TCG TTT TTG GAC TGG AT - #T AAC ACG ACA ATG         TCA      768                                                                     Tyr Thr Arg Ile Thr Ser Phe Leu Asp Trp Il - #e Asn Thr Thr Met Ser                           245  - #               250  - #               255               - - GAA AAT                - #                  - #                  -       #          774                                                                   Glu Asn                                                                         - -  - - (2) INFORMATION FOR SEQ ID NO:19:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 258 amino - #acids                                                 (B) TYPE: amino acid                                                           (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:19:                               - - Met Lys Leu Ile Ile Val Leu Ala Phe Val Le - #u Gly Ile Cys Ser         Gly                                                                                1               5 - #                 10 - #                 15              - - Ser Pro His Ser Arg Ile Ile Cys Gly Gln As - #n Ala Lys Lys Asn Ser                    20     - #             25     - #             30                   - - Ala Pro Tyr Met Ala Ser Val Gln Leu Leu As - #p Lys Val Glu Gly Val                35         - #         40         - #         45                       - - Glu Lys Leu Phe His Phe Cys Gly Gly Ala Il - #e Val Asn Asp Arg Trp            50             - #     55             - #     60                           - - Ile Leu Thr Ala Ala His Cys Leu Arg Gly Ly - #s Asp His Leu Leu Asp        65                 - # 70                 - # 75                 - # 80        - - Lys Leu Phe Ile Ala Val Gly Leu Thr Asn Le - #u Gly Glu Gly Gly Thr                        85 - #                 90 - #                 95               - - Val Tyr Pro Val Glu Lys Gly Ile Met His Gl - #u Glu Tyr Glu His Tyr                   100      - #           105      - #           110                   - - Asp Ile Val Asn Asp Ile Ala Leu Ile Lys Va - #l Lys Ser Pro Ile Glu               115          - #       120          - #       125                       - - Phe Asn Glu Lys Val Thr Thr Val Lys Leu Gl - #y Glu Asp Tyr Val Gly           130              - #   135              - #   140                           - - Gly Asp Val Gln Leu Arg Leu Thr Gly Trp Gl - #y Val Thr Thr Asn Glu       145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - Gly Ile Gly Ser Pro Ser Gln Lys Leu Gln Va - #l Met Thr Ala Lys         Ser                                                                                              165  - #               170  - #               175              - - Leu Thr Tyr Glu Asp Cys Lys Asn Ala Ile Ty - #r Lys Lys Thr Phe Glu                   180      - #           185      - #           190                   - - Ser Gln Ile Cys Ala Gln Ala Lys Lys Gly Th - #r Gly Ser Cys Lys Gly               195          - #       200          - #       205                       - - Asp Ser Gly Gly Pro Leu Val Gln Gly Asn As - #n Thr Leu Val Gly Leu           210              - #   215              - #   220                           - - Val Ser Trp Gly Met Gln Pro Cys Gly Ser Gl - #y Tyr Tyr Pro Asp Val       225                 2 - #30                 2 - #35                 2 -       #40                                                                               - - Tyr Thr Arg Ile Thr Ser Phe Leu Asp Trp Il - #e Asn Thr Thr Met         Ser                                                                                              245  - #               250  - #               255              - - Glu Asn                                                                    - -  - - (2) INFORMATION FOR SEQ ID NO:20:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 774 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (genomic)                                      - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:20:                               - - ATTTTCTGAC ATTGTCGTGT TAATCCAGTC CAAAAACGAT GTAATTCTTG TG -              #TAAACGTC     60                                                                  - - AGGATAATAA CCACTTCCAC AAGGTTGCAT ACCCCAGGAT ACTAAACCGA CC -             #AATGTATT    120                                                                  - - GTTTCCTTGG ACTAATGGAC CACCAGAATC ACCCTTACAA GATCCGGTTC CT -             #TTTTTAGC    180                                                                  - - CTGTGCACAA ATTTGGCTTT CGAAAGTCTT TTTATAAATT GCGTTTTTGC AA -             #TCCTCATA    240                                                                  - - AGTTAGTGAT TTGGCTGTCA TGACCTGTAA TTTTTGACTC GGGCTTCCGA TT -             #CCCTCATT    300                                                                  - - TGTCGTAACT CCCCATCCTG TCAATCGAAG TTGGACGTCT CCGCCAACAT AA -             #TCCTCACC    360                                                                  - - TAATTTTACA GTCGTTACTT TTTCATTGAA TTCTATCGGA GATTTGACTT TG -             #ATTAGTGC    420                                                                  - - AATATCGTTG ACTATGTCAT AATGTTCATA TTCTTCGTGC ATGATGCCTT TT -             #TCTACAGG    480                                                                  - - ATACACGGTG CCTCCTTCAC CTAAATTTGT CAGGCCGACT GCAATGAACA GT -             #TTGTCCAG    540                                                                  - - GAGGTGGTCT TTGCCTCTCA AACAATGTGC AGCAGTCAAA ATCCATCTAT CA -             #TTAACTAT    600                                                                  - - TGCTCCTCCG CAAAAATGAA ACAATTTTTC GACTCCTTCA ACTTTATCTA AA -             #AGTTGAAC    660                                                                  - - CGATGCCATG TATGGAGCCG AATTTTTTTT GGCATTTTGA CCACAAATTA TT -             #CTTGAATG    720                                                                  - - TGGAGAACCT GAACAAATTC CTAAAACAAA TGCTAGCACG ATGATGAGTT TC - #AT               774                                                                        - -  - - (2) INFORMATION FOR SEQ ID NO:21:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 711 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (genomic)                                      - -     (ix) FEATURE:                                                                   (A) NAME/KEY: CDS                                                              (B) LOCATION: 1..711                                                  - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:21:                               - - ATA ATT TGT GGT CAA AAT GCC AAA AAA AAT TC - #G GCT CCA TAC ATG GCA            48                                                                        Ile Ile Cys Gly Gln Asn Ala Lys Lys Asn Se - #r Ala Pro Tyr Met Ala              1               5 - #                 10 - #                 15               - - TCG GTT CAA CTT TTA GAT AAA GTT GAA GGA GT - #C GAA AAA TTG TTT CAT            96                                                                        Ser Val Gln Leu Leu Asp Lys Val Glu Gly Va - #l Glu Lys Leu Phe His                         20     - #             25     - #             30                   - - TTT TGC GGA GGA GCA ATA GTT AAT GAT AGA TG - #G ATT TTG ACT GCT GCA           144                                                                        Phe Cys Gly Gly Ala Ile Val Asn Asp Arg Tr - #p Ile Leu Thr Ala Ala                     35         - #         40         - #         45                       - - CAT TGT TTG AGA GGC AAA GAC CAC CTC CTG GA - #C AAA CTG TTC ATT GCA           192                                                                        His Cys Leu Arg Gly Lys Asp His Leu Leu As - #p Lys Leu Phe Ile Ala                 50             - #     55             - #     60                           - - GTC GGC CTG ACA AAT TTA GGT GAA GGA GGC AC - #C GTG TAT CCT GTA GAA           240                                                                        Val Gly Leu Thr Asn Leu Gly Glu Gly Gly Th - #r Val Tyr Pro Val Glu             65                 - # 70                 - # 75                 - # 80        - - AAA GGC ATC ATG CAC GAA GAA TAT GAA CAT TA - #T GAC ATA GTC AAC GAT           288                                                                        Lys Gly Ile Met His Glu Glu Tyr Glu His Ty - #r Asp Ile Val Asn Asp                             85 - #                 90 - #                 95               - - ATT GCA CTA ATC AAA GTC AAA TCT CCG ATA GA - #A TTC AAT GAA AAA GTA           336                                                                        Ile Ala Leu Ile Lys Val Lys Ser Pro Ile Gl - #u Phe Asn Glu Lys Val                        100      - #           105      - #           110                   - - ACG ACT GTA AAA TTA GGT GAG GAT TAT GTT GG - #C GGA GAC GTC CAA CTT           384                                                                        Thr Thr Val Lys Leu Gly Glu Asp Tyr Val Gl - #y Gly Asp Val Gln Leu                    115          - #       120          - #       125                       - - CGA TTG ACA GGA TGG GGA GTT ACG ACA AAT GA - #G GGA ATC GGA AGC CCG           432                                                                        Arg Leu Thr Gly Trp Gly Val Thr Thr Asn Gl - #u Gly Ile Gly Ser Pro                130              - #   135              - #   140                           - - AGT CAA AAA TTA CAG GTC ATG ACA GCC AAA TC - #A CTA ACT TAT GAG GAT           480                                                                        Ser Gln Lys Leu Gln Val Met Thr Ala Lys Se - #r Leu Thr Tyr Glu Asp            145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - TGC AAA AAC GCA ATT TAT AAA AAG ACT TTC GA - #A AGC CAA ATT TGT         GCA      528                                                                     Cys Lys Asn Ala Ile Tyr Lys Lys Thr Phe Gl - #u Ser Gln Ile Cys Ala                           165  - #               170  - #               175               - - CAG GCT AAA AAA GGA ACC GGA TCT TGT AAG GG - #T GAT TCT GGT GGT CCA           576                                                                        Gln Ala Lys Lys Gly Thr Gly Ser Cys Lys Gl - #y Asp Ser Gly Gly Pro                        180      - #           185      - #           190                   - - TTA GTC CAA GGA AAC AAT ACA TTG GTC GGT TT - #A GTA TCC TGG GGT ATG           624                                                                        Leu Val Gln Gly Asn Asn Thr Leu Val Gly Le - #u Val Ser Trp Gly Met                    195          - #       200          - #       205                       - - CAA CCT TGT GGA AGT GGT TAT TAT CCT GAC GT - #T TAC ACA AGA ATT ACA           672                                                                        Gln Pro Cys Gly Ser Gly Tyr Tyr Pro Asp Va - #l Tyr Thr Arg Ile Thr                210              - #   215              - #   220                           - - TCG TTT TTG GAC TGG ATT AAC ACG ACA ATG TC - #A GAA AAT                   - #    711                                                                     Ser Phe Leu Asp Trp Ile Asn Thr Thr Met Se - #r Glu Asn                        225                 2 - #30                 2 - #35                             - -  - - (2) INFORMATION FOR SEQ ID NO:22:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 237 amino - #acids                                                 (B) TYPE: amino acid                                                           (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:22:                               - - Ile Ile Cys Gly Gln Asn Ala Lys Lys Asn Se - #r Ala Pro Tyr Met Ala         1               5 - #                 10 - #                 15               - - Ser Val Gln Leu Leu Asp Lys Val Glu Gly Va - #l Glu Lys Leu Phe His                    20     - #             25     - #             30                   - - Phe Cys Gly Gly Ala Ile Val Asn Asp Arg Tr - #p Ile Leu Thr Ala Ala                35         - #         40         - #         45                       - - His Cys Leu Arg Gly Lys Asp His Leu Leu As - #p Lys Leu Phe Ile Ala            50             - #     55             - #     60                           - - Val Gly Leu Thr Asn Leu Gly Glu Gly Gly Th - #r Val Tyr Pro Val Glu        65                 - # 70                 - # 75                 - # 80        - - Lys Gly Ile Met His Glu Glu Tyr Glu His Ty - #r Asp Ile Val Asn Asp                        85 - #                 90 - #                 95               - - Ile Ala Leu Ile Lys Val Lys Ser Pro Ile Gl - #u Phe Asn Glu Lys Val                   100      - #           105      - #           110                   - - Thr Thr Val Lys Leu Gly Glu Asp Tyr Val Gl - #y Gly Asp Val Gln Leu               115          - #       120          - #       125                       - - Arg Leu Thr Gly Trp Gly Val Thr Thr Asn Gl - #u Gly Ile Gly Ser Pro           130              - #   135              - #   140                           - - Ser Gln Lys Leu Gln Val Met Thr Ala Lys Se - #r Leu Thr Tyr Glu Asp       145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - Cys Lys Asn Ala Ile Tyr Lys Lys Thr Phe Gl - #u Ser Gln Ile Cys         Ala                                                                                              165  - #               170  - #               175              - - Gln Ala Lys Lys Gly Thr Gly Ser Cys Lys Gl - #y Asp Ser Gly Gly Pro                   180      - #           185      - #           190                   - - Leu Val Gln Gly Asn Asn Thr Leu Val Gly Le - #u Val Ser Trp Gly Met               195          - #       200          - #       205                       - - Gln Pro Cys Gly Ser Gly Tyr Tyr Pro Asp Va - #l Tyr Thr Arg Ile Thr           210              - #   215              - #   220                           - - Ser Phe Leu Asp Trp Ile Asn Thr Thr Met Se - #r Glu Asn                   225                 2 - #30                 2 - #35                             - -  - - (2) INFORMATION FOR SEQ ID NO:23:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 924 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (genomic)                                      - -     (ix) FEATURE:                                                                   (A) NAME/KEY: CDS                                                              (B) LOCATION: 3..800                                                  - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #any amino acid                                          (B) LOCATION: 44                                                      - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #any amino acid                                          (B) LOCATION: 67                                                      - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #any amino acid                                          (B) LOCATION: 260                                                     - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:23:                               - - AA ATT TTA TTA CTG GTA TTA TTG GCA GTA TGC - # TTT GCT TCA GCT AAA             47                                                                           Ile Leu Leu Leu Val Leu Leu Ala Val - #Cys Phe Ala Ser Ala Lys                   1             - #  5                - #  10                - #  15         - - CGA GGT CCA CGA AAA CAT GTT CGC GAA ACA CA - #A AAA AGT CTT GCC TCT            95                                                                        Arg Gly Pro Arg Lys His Val Arg Glu Thr Gl - #n Lys Ser Leu Ala Ser                             20 - #                 25 - #                 30               - - GGG CGT ATT GTG GGT GGT GAA GCA GTG AGC AT - #T GAA NAC TAT GGA TGG           143                                                                        Gly Arg Ile Val Gly Gly Glu Ala Val Ser Il - #e Glu Xaa Tyr Gly Trp                         35     - #             40     - #             45                   - - CAA GTT TCT CTA CAA CGT TTT GGC AGT CAT TT - #C TGT GGA GGA TCT ATA           191                                                                        Gln Val Ser Leu Gln Arg Phe Gly Ser His Ph - #e Cys Gly Gly Ser Ile                     50         - #         55         - #         60                       - - ATA TCC AGT ANA TGG ATT CTT TCA GCT GCT CA - #T TGC TTT TAT GGA ACG           239                                                                        Ile Ser Ser Xaa Trp Ile Leu Ser Ala Ala Hi - #s Cys Phe Tyr Gly Thr                 65             - #     70             - #     75                           - - TTA TTT CCG ATT GGA TTC TCT GCG AGA GCC GG - #C AGC AGT ACT GTG AAT           287                                                                        Leu Phe Pro Ile Gly Phe Ser Ala Arg Ala Gl - #y Ser Ser Thr Val Asn             80                 - # 85                 - # 90                 - # 95        - - TCA GGA GGA ACT GTG CAT ACA ATT TTG TAT TG - #G TAT ATT CAT CCA AAT           335                                                                        Ser Gly Gly Thr Val His Thr Ile Leu Tyr Tr - #p Tyr Ile His Pro Asn                            100  - #               105  - #               110               - - TAT GAT TCA CAA AGT ACA GAC TTT GAT GTT TC - #T GTA GTT CGA CTA TTA           383                                                                        Tyr Asp Ser Gln Ser Thr Asp Phe Asp Val Se - #r Val Val Arg Leu Leu                        115      - #           120      - #           125                   - - TCT TCT TTA AAT TTG AAT GGA GGT TCT ATT CG - #A CCG GCT AGG TTA GTG           431                                                                        Ser Ser Leu Asn Leu Asn Gly Gly Ser Ile Ar - #g Pro Ala Arg Leu Val                    130          - #       135          - #       140                       - - GAT TCT GGA ACT GAT TTG CCA GCC GGT GAG AT - #G GTT ACA GTA ACT GGA           479                                                                        Asp Ser Gly Thr Asp Leu Pro Ala Gly Glu Me - #t Val Thr Val Thr Gly                145              - #   150              - #   155                           - - TGG GGA CGA CTT TCG GAA AAT ACT TCT GTT CC - #C TCG CCA TCA ACT CTT           527                                                                        Trp Gly Arg Leu Ser Glu Asn Thr Ser Val Pr - #o Ser Pro Ser Thr Leu            160                 1 - #65                 1 - #70                 1 -       #75                                                                               - - CAA GGA GTT ACA GTA CCA GTT GTA AGT AAT TC - #G GAA TGT CAA CAA         CAA      575                                                                     Gln Gly Val Thr Val Pro Val Val Ser Asn Se - #r Glu Cys Gln Gln Gln                           180  - #               185  - #               190               - - TTG CAA AAT CAG ACA ATC ACT GAC AAT ATG TT - #T TGT GCT GGT GAA TTA           623                                                                        Leu Gln Asn Gln Thr Ile Thr Asp Asn Met Ph - #e Cys Ala Gly Glu Leu                        195      - #           200      - #           205                   - - GAA GGA GGA AAG GAC TCT TGT CAA GGA GAC AG - #T AGT GGT CCC ATG GTT           671                                                                        Glu Gly Gly Lys Asp Ser Cys Gln Gly Asp Se - #r Ser Gly Pro Met Val                    210          - #       215          - #       220                       - - GAC AGC GAG GAT ACT CAA GTA GGA ATT GTA TC - #C TGG GGA ATA GGA TGT           719                                                                        Asp Ser Glu Asp Thr Gln Val Gly Ile Val Se - #r Trp Gly Ile Gly Cys                225              - #   230              - #   235                           - - GCT AGA CCC AAT TTA CCA GGA GTT TAT ACG CG - #A ATT GCT TCA TCG CCA           767                                                                        Ala Arg Pro Asn Leu Pro Gly Val Tyr Thr Ar - #g Ile Ala Ser Ser Pro            240                 2 - #45                 2 - #50                 2 -       #55                                                                               - - ATT AGA GAT TTC NTA AGA CGA ATA ACC GGA GT - #T TAATATTATT              TTATACATTT    820                                                                Ile Arg Asp Phe Xaa Arg Arg Ile Thr Gly Va - #l                                                260  - #               265                                      - - TTGACAAATA TGAGAACTAA TGAGAACTGT TGTNTTGCTA TAATTCTTTG CA -             #ACATTGTG    880                                                                  - - CATGAATAAA TTATGAATNT AATTGTTAAA AAAAAAAAAA AAAA   - #                       - #924                                                                      - -  - - (2) INFORMATION FOR SEQ ID NO:24:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 266 amino - #acids                                                 (B) TYPE: amino acid                                                           (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #any amino acid                                          (B) LOCATION: 44                                                      - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #any amino acid                                          (B) LOCATION: 67                                                      - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #any amino acid                                          (B) LOCATION: 260                                                     - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:24:                               - - Ile Leu Leu Leu Val Leu Leu Ala Val Cys Ph - #e Ala Ser Ala Lys Arg         1               5 - #                 10 - #                 15               - - Gly Pro Arg Lys His Val Arg Glu Thr Gln Ly - #s Ser Leu Ala Ser Gly                    20     - #             25     - #             30                   - - Arg Ile Val Gly Gly Glu Ala Val Ser Ile Gl - #u Xaa Tyr Gly Trp Gln                35         - #         40         - #         45                       - - Val Ser Leu Gln Arg Phe Gly Ser His Phe Cy - #s Gly Gly Ser Ile Ile            50             - #     55             - #     60                           - - Ser Ser Xaa Trp Ile Leu Ser Ala Ala His Cy - #s Phe Tyr Gly Thr Leu        65                 - # 70                 - # 75                 - # 80        - - Phe Pro Ile Gly Phe Ser Ala Arg Ala Gly Se - #r Ser Thr Val Asn Ser                        85 - #                 90 - #                 95               - - Gly Gly Thr Val His Thr Ile Leu Tyr Trp Ty - #r Ile His Pro Asn Tyr                   100      - #           105      - #           110                   - - Asp Ser Gln Ser Thr Asp Phe Asp Val Ser Va - #l Val Arg Leu Leu Ser               115          - #       120          - #       125                       - - Ser Leu Asn Leu Asn Gly Gly Ser Ile Arg Pr - #o Ala Arg Leu Val Asp           130              - #   135              - #   140                           - - Ser Gly Thr Asp Leu Pro Ala Gly Glu Met Va - #l Thr Val Thr Gly Trp       145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - Gly Arg Leu Ser Glu Asn Thr Ser Val Pro Se - #r Pro Ser Thr Leu         Gln                                                                                              165  - #               170  - #               175              - - Gly Val Thr Val Pro Val Val Ser Asn Ser Gl - #u Cys Gln Gln Gln Leu                   180      - #           185      - #           190                   - - Gln Asn Gln Thr Ile Thr Asp Asn Met Phe Cy - #s Ala Gly Glu Leu Glu               195          - #       200          - #       205                       - - Gly Gly Lys Asp Ser Cys Gln Gly Asp Ser Se - #r Gly Pro Met Val Asp           210              - #   215              - #   220                           - - Ser Glu Asp Thr Gln Val Gly Ile Val Ser Tr - #p Gly Ile Gly Cys Ala       225                 2 - #30                 2 - #35                 2 -       #40                                                                               - - Arg Pro Asn Leu Pro Gly Val Tyr Thr Arg Il - #e Ala Ser Ser Pro         Ile                                                                                              245  - #               250  - #               255              - - Arg Asp Phe Xaa Arg Arg Ile Thr Gly Val                                               260      - #           265                                          - -  - - (2) INFORMATION FOR SEQ ID NO:25:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 924 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (genomic)                                      - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:25:                               - - TTTTTTTTTT TTTTTTTAAC AATTANATTC ATAATTTATT CATGCACAAT GT -              #TGCAAAGA     60                                                                  - - ATTATAGCAA NACAACAGTT CTCATTAGTT CTCATATTTG TCAAAAATGT AT -             #AAAATAAT    120                                                                  - - ATTAAACTCC GGTTATTCGT CTTANGAAAT CTCTAATTGG CGATGAAGCA AT -             #TCGCGTAT    180                                                                  - - AAACTCCTGG TAAATTGGGT CTAGCACATC CTATTCCCCA GGATACAATT CC -             #TACTTGAG    240                                                                  - - TATCCTCGCT GTCAACCATG GGACCACTAC TGTCTCCTTG ACAAGAGTCC TT -             #TCCTCCTT    300                                                                  - - CTAATTCACC AGCACAAAAC ATATTGTCAG TGATTGTCTG ATTTTGCAAT TG -             #TTGTTGAC    360                                                                  - - ATTCCGAATT ACTTACAACT GGTACTGTAA CTCCTTGAAG AGTTGATGGC GA -             #GGGAACAG    420                                                                  - - AAGTATTTTC CGAAAGTCGT CCCCATCCAG TTACTGTAAC CATCTCACCG GC -             #TGGCAAAT    480                                                                  - - CAGTTCCAGA ATCCACTAAC CTAGCCGGTC GAATAGAACC TCCATTCAAA TT -             #TAAAGAAG    540                                                                  - - ATAATAGTCG AACTACAGAA ACATCAAAGT CTGTACTTTG TGAATCATAA TT -             #TGGATGAA    600                                                                  - - TATACCAATA CAAAATTGTA TGCACAGTTC CTCCTGAATT CACAGTACTG CT -             #GCCGGCTC    660                                                                  - - TCGCAGAGAA TCCAATCGGA AATAACGTTC CATAAAAGCA ATGAGCAGCT GA -             #AAGAATCC    720                                                                  - - ATNTACTGGA TATTATAGAT CCTCCACAGA AATGACTGCC AAAACGTTGT AG -             #AGAAACTT    780                                                                  - - GCCATCCATA GTNTTCAATG CTCACTGCTT CACCACCCAC AATACGCCCA GA -             #GGCAAGAC    840                                                                  - - TTTTTTGTGT TTCGCGAACA TGTTTTCGTG GACCTNGTTT AGCTGAAGCA AA -             #GCATACTG    900                                                                  - - CCAATAATAC CAGTAATAAA ATTT          - #                  - #                    924                                                                      - -  - - (2) INFORMATION FOR SEQ ID NO:26:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 699 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (genomic)                                      - -     (ix) FEATURE:                                                                   (A) NAME/KEY: CDS                                                              (B) LOCATION: 1..699                                                  - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #any amino acid                                          (B) LOCATION: 11                                                      - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #any amino acid                                          (B) LOCATION: 34                                                      - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #any amino acid                                          (B) LOCATION: 227                                                     - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:26:                               - - ATT GTG GGT GGT GAA GCA GTG AGC ATT GAA NA - #C TAT GGA TGG CAA GTT            48                                                                        Ile Val Gly Gly Glu Ala Val Ser Ile Glu Xa - #a Tyr Gly Trp Gln Val              1               5 - #                 10 - #                 15               - - TCT CTA CAA CGT TTT GGC AGT CAT TTC TGT GG - #A GGA TCT ATA ATA TCC            96                                                                        Ser Leu Gln Arg Phe Gly Ser His Phe Cys Gl - #y Gly Ser Ile Ile Ser                         20     - #             25     - #             30                   - - AGT ANA TGG ATT CTT TCA GCT GCT CAT TGC TT - #T TAT GGA ACG TTA TTT           144                                                                        Ser Xaa Trp Ile Leu Ser Ala Ala His Cys Ph - #e Tyr Gly Thr Leu Phe                     35         - #         40         - #         45                       - - CCG ATT GGA TTC TCT GCG AGA GCC GGC AGC AG - #T ACT GTG AAT TCA GGA           192                                                                        Pro Ile Gly Phe Ser Ala Arg Ala Gly Ser Se - #r Thr Val Asn Ser Gly                 50             - #     55             - #     60                           - - GGA ACT GTG CAT ACA ATT TTG TAT TGG TAT AT - #T CAT CCA AAT TAT GAT           240                                                                        Gly Thr Val His Thr Ile Leu Tyr Trp Tyr Il - #e His Pro Asn Tyr Asp             65                 - # 70                 - # 75                 - # 80        - - TCA CAA AGT ACA GAC TTT GAT GTT TCT GTA GT - #T CGA CTA TTA TCT TCT           288                                                                        Ser Gln Ser Thr Asp Phe Asp Val Ser Val Va - #l Arg Leu Leu Ser Ser                             85 - #                 90 - #                 95               - - TTA AAT TTG AAT GGA GGT TCT ATT CGA CCG GC - #T AGG TTA GTG GAT TCT           336                                                                        Leu Asn Leu Asn Gly Gly Ser Ile Arg Pro Al - #a Arg Leu Val Asp Ser                        100      - #           105      - #           110                   - - GGA ACT GAT TTG CCA GCC GGT GAG ATG GTT AC - #A GTA ACT GGA TGG GGA           384                                                                        Gly Thr Asp Leu Pro Ala Gly Glu Met Val Th - #r Val Thr Gly Trp Gly                    115          - #       120          - #       125                       - - CGA CTT TCG GAA AAT ACT TCT GTT CCC TCG CC - #A TCA ACT CTT CAA GGA           432                                                                        Arg Leu Ser Glu Asn Thr Ser Val Pro Ser Pr - #o Ser Thr Leu Gln Gly                130              - #   135              - #   140                           - - GTT ACA GTA CCA GTT GTA AGT AAT TCG GAA TG - #T CAA CAA CAA TTG CAA           480                                                                        Val Thr Val Pro Val Val Ser Asn Ser Glu Cy - #s Gln Gln Gln Leu Gln            145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - AAT CAG ACA ATC ACT GAC AAT ATG TTT TGT GC - #T GGT GAA TTA GAA         GGA      528                                                                     Asn Gln Thr Ile Thr Asp Asn Met Phe Cys Al - #a Gly Glu Leu Glu Gly                           165  - #               170  - #               175               - - GGA AAG GAC TCT TGT CAA GGA GAC AGT AGT GG - #T CCC ATG GTT GAC AGC           576                                                                        Gly Lys Asp Ser Cys Gln Gly Asp Ser Ser Gl - #y Pro Met Val Asp Ser                        180      - #           185      - #           190                   - - GAG GAT ACT CAA GTA GGA ATT GTA TCC TGG GG - #A ATA GGA TGT GCT AGA           624                                                                        Glu Asp Thr Gln Val Gly Ile Val Ser Trp Gl - #y Ile Gly Cys Ala Arg                    195          - #       200          - #       205                       - - CCC AAT TTA CCA GGA GTT TAT ACG CGA ATT GC - #T TCA TCG CCA ATT AGA           672                                                                        Pro Asn Leu Pro Gly Val Tyr Thr Arg Ile Al - #a Ser Ser Pro Ile Arg                210              - #   215              - #   220                           - - GAT TTC NTA AGA CGA ATA ACC GGA GTT    - #                  - #                 699                                                                      Asp Phe Xaa Arg Arg Ile Thr Gly Val                                            225                 2 - #30                                                     - -  - - (2) INFORMATION FOR SEQ ID NO:27:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 233 amino - #acids                                                 (B) TYPE: amino acid                                                           (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #any amino acid                                          (B) LOCATION: 11                                                      - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #any amino acid                                          (B) LOCATION: 34                                                      - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #any amino acid                                          (B) LOCATION: 227                                                     - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:27:                               - - Ile Val Gly Gly Glu Ala Val Ser Ile Glu Xa - #a Tyr Gly Trp Gln Val         1               5 - #                 10 - #                 15               - - Ser Leu Gln Arg Phe Gly Ser His Phe Cys Gl - #y Gly Ser Ile Ile Ser                    20     - #             25     - #             30                   - - Ser Xaa Trp Ile Leu Ser Ala Ala His Cys Ph - #e Tyr Gly Thr Leu Phe                35         - #         40         - #         45                       - - Pro Ile Gly Phe Ser Ala Arg Ala Gly Ser Se - #r Thr Val Asn Ser Gly            50             - #     55             - #     60                           - - Gly Thr Val His Thr Ile Leu Tyr Trp Tyr Il - #e His Pro Asn Tyr Asp        65                 - # 70                 - # 75                 - # 80        - - Ser Gln Ser Thr Asp Phe Asp Val Ser Val Va - #l Arg Leu Leu Ser Ser                        85 - #                 90 - #                 95               - - Leu Asn Leu Asn Gly Gly Ser Ile Arg Pro Al - #a Arg Leu Val Asp Ser                   100      - #           105      - #           110                   - - Gly Thr Asp Leu Pro Ala Gly Glu Met Val Th - #r Val Thr Gly Trp Gly               115          - #       120          - #       125                       - - Arg Leu Ser Glu Asn Thr Ser Val Pro Ser Pr - #o Ser Thr Leu Gln Gly           130              - #   135              - #   140                           - - Val Thr Val Pro Val Val Ser Asn Ser Glu Cy - #s Gln Gln Gln Leu Gln       145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - Asn Gln Thr Ile Thr Asp Asn Met Phe Cys Al - #a Gly Glu Leu Glu         Gly                                                                                              165  - #               170  - #               175              - - Gly Lys Asp Ser Cys Gln Gly Asp Ser Ser Gl - #y Pro Met Val Asp Ser                   180      - #           185      - #           190                   - - Glu Asp Thr Gln Val Gly Ile Val Ser Trp Gl - #y Ile Gly Cys Ala Arg               195          - #       200          - #       205                       - - Pro Asn Leu Pro Gly Val Tyr Thr Arg Ile Al - #a Ser Ser Pro Ile Arg           210              - #   215              - #   220                           - - Asp Phe Xaa Arg Arg Ile Thr Gly Val                                       225                 2 - #30                                                     - -  - - (2) INFORMATION FOR SEQ ID NO:28:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 699 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (genomic)                                      - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:28:                               - - AACTCCGGTT ATTCGTCTTA NGAAATCTCT AATTGGCGAT GAAGCAATTC GC -              #GTATAAAC     60                                                                  - - TCCTGGTAAA TTGGGTCTAG CACATCCTAT TCCCCAGGAT ACAATTCCTA CT -             #TGAGTATC    120                                                                  - - CTCGCTGTCA ACCATGGGAC CACTACTGTC TCCTTGACAA GAGTCCTTTC CT -             #CCTTCTAA    180                                                                  - - TTCACCAGCA CAAAACATAT TGTCAGTGAT TGTCTGATTT TGCAATTGTT GT -             #TGACATTC    240                                                                  - - CGAATTACTT ACAACTGGTA CTGTAACTCC TTGAAGAGTT GATGGCGAGG GA -             #ACAGAAGT    300                                                                  - - ATTTTCCGAA AGTCGTCCCC ATCCAGTTAC TGTAACCATC TCACCGGCTG GC -             #AAATCAGT    360                                                                  - - TCCAGAATCC ACTAACCTAG CCGGTCGAAT AGAACCTCCA TTCAAATTTA AA -             #GAAGATAA    420                                                                  - - TAGTCGAACT ACAGAAACAT CAAAGTCTGT ACTTTGTGAA TCATAATTTG GA -             #TGAATATA    480                                                                  - - CCAATACAAA ATTGTATGCA CAGTTCCTCC TGAATTCACA GTACTGCTGC CG -             #GCTCTCGC    540                                                                  - - AGAGAATCCA ATCGGAAATA ACGTTCCATA AAAGCAATGA GCAGCTGAAA GA -             #ATCCATNT    600                                                                  - - ACTGGATATT ATAGATCCTC CACAGAAATG ACTGCCAAAA CGTTGTAGAG AA -             #ACTTGCCA    660                                                                  - - TCCATAGTNT TCAATGCTCA CTGCTTCACC ACCCACAAT      - #                       - #   699                                                                      - -  - - (2) INFORMATION FOR SEQ ID NO:29:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 1894 base - #pairs                                                 (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (genomic)                                      - -     (ix) FEATURE:                                                                   (A) NAME/KEY: CDS                                                              (B) LOCATION: 335..1535                                               - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:29:                               - - ACGCGACGGG CAGTCTCTTT CAGACCGCGG CCGAACGATG TTTTTGACGG TT -              #TAATTTCA     60                                                                  - - ATTTTTGCAA CTTGTGACTT CGAATACACC CGTACGTGAC CTATCCACTT TA -             #CCATCGGC    120                                                                  - - CGACCGTGAA AGTTGTGTTT GTGCTATTGA AAATTTCGTG CTCAATAATA AA -             #TATTCAGT    180                                                                  - - TTTTTTGTCT AATCAGGATA TTTAAATTTA TGTGTACAAG TGTTAAACGC AA -             #TCTCGTCG    240                                                                  - - CTCTTCCTAA TTTTCTTGTT GCCAATDCTG GCCTCAGTGC ATCGGCCGAC GC -             #AAATGTGC    300                                                                  - - TTGCAAAATA GAGAAATCCG GTGAAATCAC ACAT ATG TTA GCG A - #TC GTC CCG             352                                                                                         - #                  - #  Met Leu Ala Ile Val Pro                              - #                  - #    1              - # 5              - - TCA AAC GGA GCG TTC GCA GAC CAT GCC AAC CT - #T GGT GGA GTT GAT GGT           400                                                                        Ser Asn Gly Ala Phe Ala Asp His Ala Asn Le - #u Gly Gly Val Asp Gly                         10     - #             15     - #             20                   - - CTT TCT GGT TTG ATT CTG GTC GCT GTT GCG AT - #A TCT TCG ATT GGA TAT           448                                                                        Leu Ser Gly Leu Ile Leu Val Ala Val Ala Il - #e Ser Ser Ile Gly Tyr                     25         - #         30         - #         35                       - - GCG GAC GCG GCG AAC GTT GCG CAG GAC GGA CA - #T CCG TCC AGC CAG CAA           496                                                                        Ala Asp Ala Ala Asn Val Ala Gln Asp Gly Hi - #s Pro Ser Ser Gln Gln                 40             - #     45             - #     50                           - - GAG CAG GAG ATC CTG CTG CTG AAT GCC TTA GC - #T CGC AGG AAC GGA GCG           544                                                                        Glu Gln Glu Ile Leu Leu Leu Asn Ala Leu Al - #a Arg Arg Asn Gly Ala             55                 - # 60                 - # 65                 - # 70        - - ACG GGG CAC CAA TTT GAC GTA GAT CAA GAT TC - #A ATT ATG GAT ATG CTA           592                                                                        Thr Gly His Gln Phe Asp Val Asp Gln Asp Se - #r Ile Met Asp Met Leu                             75 - #                 80 - #                 85               - - GGA AGA ATG ATA CCT CAG ACT TGC CGG TAC AA - #A GGC GAA CGG TTC GAG           640                                                                        Gly Arg Met Ile Pro Gln Thr Cys Arg Tyr Ly - #s Gly Glu Arg Phe Glu                         90     - #             95     - #            100                   - - TGC GGT TTG TCA ATT TCG TGC GTC CTG GGC GG - #C GGA AAA CCT CTT GAC           688                                                                        Cys Gly Leu Ser Ile Ser Cys Val Leu Gly Gl - #y Gly Lys Pro Leu Asp                    105          - #       110          - #       115                       - - CTG TGC AGC GGC GGA ATG ATC TGG TCG TGC TG - #C GTC GAC AGG GAC ATT           736                                                                        Leu Cys Ser Gly Gly Met Ile Trp Ser Cys Cy - #s Val Asp Arg Asp Ile                120              - #   125              - #   130                           - - CGG CCT GAG CCG CAG CAC CAG GGC GCT CTG CA - #G AAC GCA ACT TGT GGA           784                                                                        Arg Pro Glu Pro Gln His Gln Gly Ala Leu Gl - #n Asn Ala Thr Cys Gly            135                 1 - #40                 1 - #45                 1 -       #50                                                                               - - GAA TTG TAC ACG AGG TCT AAT AGA ATC GTA GG - #A GGT CAT TCA ACA         GGA      832                                                                     Glu Leu Tyr Thr Arg Ser Asn Arg Ile Val Gl - #y Gly His Ser Thr Gly                           155  - #               160  - #               165               - - TTC GGG TCT CAT CCT TGG CAG GCG GCT TTG AT - #C AAA TCA GGA TTT TTG           880                                                                        Phe Gly Ser His Pro Trp Gln Ala Ala Leu Il - #e Lys Ser Gly Phe Leu                        170      - #           175      - #           180                   - - AGT AAA AAA TTA TCT TGC GGT GGC GCT TTA GT - #T AGC GAT CGA TGG GTT           928                                                                        Ser Lys Lys Leu Ser Cys Gly Gly Ala Leu Va - #l Ser Asp Arg Trp Val                    185          - #       190          - #       195                       - - ATA ACT GCT GCA CAT TGC GTT GCC ACG ACA CC - #A AAT TCG AAC CTG AAG           976                                                                        Ile Thr Ala Ala His Cys Val Ala Thr Thr Pr - #o Asn Ser Asn Leu Lys                200              - #   205              - #   210                           - - GTG CGA TTG GGC GAA TGG GAC GTC CGC GAC CA - #C GAT GAG CGA CTG AAC          1024                                                                        Val Arg Leu Gly Glu Trp Asp Val Arg Asp Hi - #s Asp Glu Arg Leu Asn            215                 2 - #20                 2 - #25                 2 -       #30                                                                               - - CAC GAG GAA TAC GCA ATC GAA CGC AAA GAA GT - #T CAT CCT TCA TAT         TCA     1072                                                                     His Glu Glu Tyr Ala Ile Glu Arg Lys Glu Va - #l His Pro Ser Tyr Ser                           235  - #               240  - #               245               - - CCA ACC GAT TTC CGG AAT GAT GTA GCC TTA GT - #G AAA CTC GAT AGA ACT          1120                                                                        Pro Thr Asp Phe Arg Asn Asp Val Ala Leu Va - #l Lys Leu Asp Arg Thr                        250      - #           255      - #           260                   - - GTT ATT TTC AAA CAA CAT ATT TTA CCT GTC TG - #C TTA CCT CAT AAG CAA          1168                                                                        Val Ile Phe Lys Gln His Ile Leu Pro Val Cy - #s Leu Pro His Lys Gln                    265          - #       270          - #       275                       - - ATG AAA CTG GCT GGA AAA ATG GCA ACA GTC GC - #C GGA TGG GGA CGG ACG          1216                                                                        Met Lys Leu Ala Gly Lys Met Ala Thr Val Al - #a Gly Trp Gly Arg Thr                280              - #   285              - #   290                           - - AGG CAC GGG CAG AGC ACT GTG CCG GCT GTC TT - #A CAA GAA GTC GAT GTC          1264                                                                        Arg His Gly Gln Ser Thr Val Pro Ala Val Le - #u Gln Glu Val Asp Val            295                 3 - #00                 3 - #05                 3 -       #10                                                                               - - GAG GTG ATT CCG AAT GAA AGA TGC CAG AGG TG - #G TTC CGT GCT GCG         GGT     1312                                                                     Glu Val Ile Pro Asn Glu Arg Cys Gln Arg Tr - #p Phe Arg Ala Ala Gly                           315  - #               320  - #               325               - - CGA CGA GAA ACC ATT CAC GAT GTC TTT CTC TG - #C GCC GGA TAT AAA GAG          1360                                                                        Arg Arg Glu Thr Ile His Asp Val Phe Leu Cy - #s Ala Gly Tyr Lys Glu                        330      - #           335      - #           340                   - - GGT GGT CGT GAT TCA TGC CAA GGT GAT TCT GG - #A GGT CCT CTA ATA ATG          1408                                                                        Gly Gly Arg Asp Ser Cys Gln Gly Asp Ser Gl - #y Gly Pro Leu Ile Met                    345          - #       350          - #       355                       - - CAG ATT GAG GGT AGA AGG ACC CTT GTG GGT CT - #A GTT TCT TGG GGC ATT          1456                                                                        Gln Ile Glu Gly Arg Arg Thr Leu Val Gly Le - #u Val Ser Trp Gly Ile                360              - #   365              - #   370                           - - GGA TGT GGT CGT GAG CAT TTA CCA GGC GTA TA - #T ACC AAT ATA CAA AAA          1504                                                                        Gly Cys Gly Arg Glu His Leu Pro Gly Val Ty - #r Thr Asn Ile Gln Lys            375                 3 - #80                 3 - #85                 3 -       #90                                                                               - - TTC ATA CCG TGG ATC GAC AAA GTA ATG GGA T - #AATTTTTATT CCATCGAGCT          1555                                                                        Phe Ile Pro Trp Ile Asp Lys Val Met Gly                                                        395  - #               400                                      - - TACCCAAAGT ATTTATTAAG TGTTAATCGA AAGTTCCAAT AATAAATTAA TT -              #TAAAATTC   1615                                                                  - - TAAAGACGGG AATTTGAAAG ACCAAAAAGA CATACTTGTG ATTGTGTAAT TT -             #TTATGATT   1675                                                                  - - AACTTTACAT CATCTGTGCT TAATTATTAA TTTGTATTAT TCTTGCAAAT AT -             #TTCAAGAG   1735                                                                  - - TTACCGAAAA GTTTGCTAAT CGATAATGAT ATTTTAAGAA AAACAACTGC TG -             #CTGATTCA   1795                                                                  - - GTCAATGTTA GAATAATTAT GTTTACTAAA TAATATTAAG TTCTGATTAG TA -             #AATAAATA   1855                                                                  - - GCAAAATTAT CTAAATATAT ATAAAAAAAA AAAAAAAAA      - #                       - #  1894                                                                      - -  - - (2) INFORMATION FOR SEQ ID NO:30:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 400 amino - #acids                                                 (B) TYPE: amino acid                                                           (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:30:                               - - Met Leu Ala Ile Val Pro Ser Asn Gly Ala Ph - #e Ala Asp His Ala Asn         1               5 - #                 10 - #                 15               - - Leu Gly Gly Val Asp Gly Leu Ser Gly Leu Il - #e Leu Val Ala Val Ala                    20     - #             25     - #             30                   - - Ile Ser Ser Ile Gly Tyr Ala Asp Ala Ala As - #n Val Ala Gln Asp Gly                35         - #         40         - #         45                       - - His Pro Ser Ser Gln Gln Glu Gln Glu Ile Le - #u Leu Leu Asn Ala Leu            50             - #     55             - #     60                           - - Ala Arg Arg Asn Gly Ala Thr Gly His Gln Ph - #e Asp Val Asp Gln Asp        65                 - # 70                 - # 75                 - # 80        - - Ser Ile Met Asp Met Leu Gly Arg Met Ile Pr - #o Gln Thr Cys Arg Tyr                        85 - #                 90 - #                 95               - - Lys Gly Glu Arg Phe Glu Cys Gly Leu Ser Il - #e Ser Cys Val Leu Gly                   100      - #           105      - #           110                   - - Gly Gly Lys Pro Leu Asp Leu Cys Ser Gly Gl - #y Met Ile Trp Ser Cys               115          - #       120          - #       125                       - - Cys Val Asp Arg Asp Ile Arg Pro Glu Pro Gl - #n His Gln Gly Ala Leu           130              - #   135              - #   140                           - - Gln Asn Ala Thr Cys Gly Glu Leu Tyr Thr Ar - #g Ser Asn Arg Ile Val       145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - Gly Gly His Ser Thr Gly Phe Gly Ser His Pr - #o Trp Gln Ala Ala         Leu                                                                                              165  - #               170  - #               175              - - Ile Lys Ser Gly Phe Leu Ser Lys Lys Leu Se - #r Cys Gly Gly Ala Leu                   180      - #           185      - #           190                   - - Val Ser Asp Arg Trp Val Ile Thr Ala Ala Hi - #s Cys Val Ala Thr Thr               195          - #       200          - #       205                       - - Pro Asn Ser Asn Leu Lys Val Arg Leu Gly Gl - #u Trp Asp Val Arg Asp           210              - #   215              - #   220                           - - His Asp Glu Arg Leu Asn His Glu Glu Tyr Al - #a Ile Glu Arg Lys Glu       225                 2 - #30                 2 - #35                 2 -       #40                                                                               - - Val His Pro Ser Tyr Ser Pro Thr Asp Phe Ar - #g Asn Asp Val Ala         Leu                                                                                              245  - #               250  - #               255              - - Val Lys Leu Asp Arg Thr Val Ile Phe Lys Gl - #n His Ile Leu Pro Val                   260      - #           265      - #           270                   - - Cys Leu Pro His Lys Gln Met Lys Leu Ala Gl - #y Lys Met Ala Thr Val               275          - #       280          - #       285                       - - Ala Gly Trp Gly Arg Thr Arg His Gly Gln Se - #r Thr Val Pro Ala Val           290              - #   295              - #   300                           - - Leu Gln Glu Val Asp Val Glu Val Ile Pro As - #n Glu Arg Cys Gln Arg       305                 3 - #10                 3 - #15                 3 -       #20                                                                               - - Trp Phe Arg Ala Ala Gly Arg Arg Glu Thr Il - #e His Asp Val Phe         Leu                                                                                              325  - #               330  - #               335              - - Cys Ala Gly Tyr Lys Glu Gly Gly Arg Asp Se - #r Cys Gln Gly Asp Ser                   340      - #           345      - #           350                   - - Gly Gly Pro Leu Ile Met Gln Ile Glu Gly Ar - #g Arg Thr Leu Val Gly               355          - #       360          - #       365                       - - Leu Val Ser Trp Gly Ile Gly Cys Gly Arg Gl - #u His Leu Pro Gly Val           370              - #   375              - #   380                           - - Tyr Thr Asn Ile Gln Lys Phe Ile Pro Trp Il - #e Asp Lys Val Met Gly       385                 3 - #90                 3 - #95                 4 -       #00                                                                               - -  - - (2) INFORMATION FOR SEQ ID NO:31:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 1894 base - #pairs                                                 (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (genomic)                                      - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:31:                               - - TTTTTTTTTT TTTTTTTATA TATATTTAGA TAATTTTGCT ATTTATTTAC TA -             #ATCAGAAC     60                                                                  - - TTAATATTAT TTAGTAAACA TAATTATTCT AACATTGACT GAATCAGCAG CA -             #GTTGTTTT    120                                                                  - - TCTTAAAATA TCATTATCGA TTAGCAAACT TTTCGGTAAC TCTTGAAATA TT -             #TGCAAGAA    180                                                                  - - TAATACAAAT TAATAATTAA GCACAGATGA TGTAAAGTTA ATCATAAAAA TT -             #ACACAATC    240                                                                  - - ACAAGTATGT CTTTTTGGTC TTTCAAATTC CCGTCTTTAG AATTTTAAAT TA -             #ATTTATTA    300                                                                  - - TTGGAACTTT CGATTAACAC TTAATAAATA CTTTGGGTAA GCTCGATGGA AT -             #AAAAATTA    360                                                                  - - TCCCATTACT TTGTCGATCC ACGGTATGAA TTTTTGTATA TTGGTATATA CG -             #CCTGGTAA    420                                                                  - - ATGCTCACGA CCACATCCAA TGCCCCAAGA AACTAGACCC ACAAGGGTCC TT -             #CTACCCTC    480                                                                  - - AATCTGCATT ATTAGAGGAC CTCCAGAATC ACCTTGGCAT GAATCACGAC CA -             #CCCTCTTT    540                                                                  - - ATATCCGGCG CAGAGAAAGA CATCGTGAAT GGTTTCTCGT CGACCCGCAG CA -             #CGGAACCA    600                                                                  - - CCTCTGGCAT CTTTCATTCG GAATCACCTC GACATCGACT TCTTGTAAGA CA -             #GCCGGCAC    660                                                                  - - AGTGCTCTGC CCGTGCCTCG TCCGTCCCCA TCCGGCGACT GTTGCCATTT TT -             #CCAGCCAG    720                                                                  - - TTTCATTTGC TTATGAGGTA AGCAGACAGG TAAAATATGT TGTTTGAAAA TA -             #ACAGTTCT    780                                                                  - - ATCGAGTTTC ACTAAGGCTA CATCATTCCG GAAATCGGTT GGTGAATATG AA -             #GGATGAAC    840                                                                  - - TTCTTTGCGT TCGATTGCGT ATTCCTCGTG GTTCAGTCGC TCATCGTGGT CG -             #CGGACGTC    900                                                                  - - CCATTCGCCC AATCGCACCT TCAGGTTCGA ATTTGGTGTC GTGGCAACGC AA -             #TGTGCAGC    960                                                                  - - AGTTATAACC CATCGATCGC TAACTAAAGC GCCACCGCAA GATAATTTTT TA -             #CTCAAAAA   1020                                                                  - - TCCTGATTTG ATCAAAGCCG CCTGCCAAGG ATGAGACCCG AATCCTGTTG AA -             #TGACCTCC   1080                                                                  - - TACGATTCTA TTAGACCTCG TGTACAATTC TCCACAAGTT GCGTTCTGCA GA -             #GCGCCCTG   1140                                                                  - - GTGCTGCGGC TCAGGCCGAA TGTCCCTGTC GACGCAGCAC GACCAGATCA TT -             #CCGCCGCT   1200                                                                  - - GCACAGGTCA AGAGGTTTTC CGCCGCCCAG GACGCACGAA ATTGACAAAC CG -             #CACTCGAA   1260                                                                  - - CCGTTCGCCT TTGTACCGGC AAGTCTGAGG TATCATTCTT CCTAGCATAT CC -             #ATAATTGA   1320                                                                  - - ATCTTGATCT ACGTCAAATT GGTGCCCCGT CGCTCCGTTC CTGCGAGCTA AG -             #GCATTCAG   1380                                                                  - - CAGCAGGATC TCCTGCTCTT GCTGGCTGGA CGGATGTCCG TCCTGCGCAA CG -             #TTCGCCGC   1440                                                                  - - GTCCGCATAT CCAATCGAAG ATATCGCAAC AGCGACCAGA ATCAAACCAG AA -             #AGACCATC   1500                                                                  - - AACTCCACCA AGGTTGGCAT GGTCTGCGAA CGCTCCGTTT GACGGGACGA TC -             #GCTAACAT   1560                                                                  - - ATGTGTGATT TCACCGGATT TCTCTATTTT GCAAGCACAT TTGCGTCGGC CG -             #ATGCACTG   1620                                                                  - - AGGCCAGHAT TGGCAACAAG AAAATTAGGA AGAGCGACGA GATTGCGTTT AA -             #CACTTGTA   1680                                                                  - - CACATAAATT TAAATATCCT GATTAGACAA AAAAACTGAA TATTTATTAT TG -             #AGCACGAA   1740                                                                  - - ATTTTCAATA GCACAAACAC AACTTTCACG GTCGGCCGAT GGTAAAGTGG AT -             #AGGTCACG   1800                                                                  - - TACGGGTGTA TTCGAAGTCA CAAGTTGCAA AAATTGAAAT TAAACCGTCA AA -             #AACATCGT   1860                                                                  - - TCGGCCGCGG TCTGAAAGAG ACTGCCCGTC GCGT       - #                  -      #      1894                                                                      - -  - - (2) INFORMATION FOR SEQ ID NO:32:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 1200 base - #pairs                                                 (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (genomic)                                      - -     (ix) FEATURE:                                                                   (A) NAME/KEY: CDS                                                              (B) LOCATION: 1..1200                                                 - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:32:                               - - ATG TTA GCG ATC GTC CCG TCA AAC GGA GCG TT - #C GCA GAC CAT GCC AAC            48                                                                        Met Leu Ala Ile Val Pro Ser Asn Gly Ala Ph - #e Ala Asp His Ala Asn              1               5 - #                 10 - #                 15               - - CTT GGT GGA GTT GAT GGT CTT TCT GGT TTG AT - #T CTG GTC GCT GTT GCG            96                                                                        Leu Gly Gly Val Asp Gly Leu Ser Gly Leu Il - #e Leu Val Ala Val Ala                         20     - #             25     - #             30                   - - ATA TCT TCG ATT GGA TAT GCG GAC GCG GCG AA - #C GTT GCG CAG GAC GGA           144                                                                        Ile Ser Ser Ile Gly Tyr Ala Asp Ala Ala As - #n Val Ala Gln Asp Gly                     35         - #         40         - #         45                       - - CAT CCG TCC AGC CAG CAA GAG CAG GAG ATC CT - #G CTG CTG AAT GCC TTA           192                                                                        His Pro Ser Ser Gln Gln Glu Gln Glu Ile Le - #u Leu Leu Asn Ala Leu                 50             - #     55             - #     60                           - - GCT CGC AGG AAC GGA GCG ACG GGG CAC CAA TT - #T GAC GTA GAT CAA GAT           240                                                                        Ala Arg Arg Asn Gly Ala Thr Gly His Gln Ph - #e Asp Val Asp Gln Asp             65                 - # 70                 - # 75                 - # 80        - - TCA ATT ATG GAT ATG CTA GGA AGA ATG ATA CC - #T CAG ACT TGC CGG TAC           288                                                                        Ser Ile Met Asp Met Leu Gly Arg Met Ile Pr - #o Gln Thr Cys Arg Tyr                             85 - #                 90 - #                 95               - - AAA GGC GAA CGG TTC GAG TGC GGT TTG TCA AT - #T TCG TGC GTC CTG GGC           336                                                                        Lys Gly Glu Arg Phe Glu Cys Gly Leu Ser Il - #e Ser Cys Val Leu Gly                        100      - #           105      - #           110                   - - GGC GGA AAA CCT CTT GAC CTG TGC AGC GGC GG - #A ATG ATC TGG TCG TGC           384                                                                        Gly Gly Lys Pro Leu Asp Leu Cys Ser Gly Gl - #y Met Ile Trp Ser Cys                    115          - #       120          - #       125                       - - TGC GTC GAC AGG GAC ATT CGG CCT GAG CCG CA - #G CAC CAG GGC GCT CTG           432                                                                        Cys Val Asp Arg Asp Ile Arg Pro Glu Pro Gl - #n His Gln Gly Ala Leu                130              - #   135              - #   140                           - - CAG AAC GCA ACT TGT GGA GAA TTG TAC ACG AG - #G TCT AAT AGA ATC GTA           480                                                                        Gln Asn Ala Thr Cys Gly Glu Leu Tyr Thr Ar - #g Ser Asn Arg Ile Val            145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - GGA GGT CAT TCA ACA GGA TTC GGG TCT CAT CC - #T TGG CAG GCG GCT         TTG      528                                                                     Gly Gly His Ser Thr Gly Phe Gly Ser His Pr - #o Trp Gln Ala Ala Leu                           165  - #               170  - #               175               - - ATC AAA TCA GGA TTT TTG AGT AAA AAA TTA TC - #T TGC GGT GGC GCT TTA           576                                                                        Ile Lys Ser Gly Phe Leu Ser Lys Lys Leu Se - #r Cys Gly Gly Ala Leu                        180      - #           185      - #           190                   - - GTT AGC GAT CGA TGG GTT ATA ACT GCT GCA CA - #T TGC GTT GCC ACG ACA           624                                                                        Val Ser Asp Arg Trp Val Ile Thr Ala Ala Hi - #s Cys Val Ala Thr Thr                    195          - #       200          - #       205                       - - CCA AAT TCG AAC CTG AAG GTG CGA TTG GGC GA - #A TGG GAC GTC CGC GAC           672                                                                        Pro Asn Ser Asn Leu Lys Val Arg Leu Gly Gl - #u Trp Asp Val Arg Asp                210              - #   215              - #   220                           - - CAC GAT GAG CGA CTG AAC CAC GAG GAA TAC GC - #A ATC GAA CGC AAA GAA           720                                                                        His Asp Glu Arg Leu Asn His Glu Glu Tyr Al - #a Ile Glu Arg Lys Glu            225                 2 - #30                 2 - #35                 2 -       #40                                                                               - - GTT CAT CCT TCA TAT TCA CCA ACC GAT TTC CG - #G AAT GAT GTA GCC         TTA      768                                                                     Val His Pro Ser Tyr Ser Pro Thr Asp Phe Ar - #g Asn Asp Val Ala Leu                           245  - #               250  - #               255               - - GTG AAA CTC GAT AGA ACT GTT ATT TTC AAA CA - #A CAT ATT TTA CCT GTC           816                                                                        Val Lys Leu Asp Arg Thr Val Ile Phe Lys Gl - #n His Ile Leu Pro Val                        260      - #           265      - #           270                   - - TGC TTA CCT CAT AAG CAA ATG AAA CTG GCT GG - #A AAA ATG GCA ACA GTC           864                                                                        Cys Leu Pro His Lys Gln Met Lys Leu Ala Gl - #y Lys Met Ala Thr Val                    275          - #       280          - #       285                       - - GCC GGA TGG GGA CGG ACG AGG CAC GGG CAG AG - #C ACT GTG CCG GCT GTC           912                                                                        Ala Gly Trp Gly Arg Thr Arg His Gly Gln Se - #r Thr Val Pro Ala Val                290              - #   295              - #   300                           - - TTA CAA GAA GTC GAT GTC GAG GTG ATT CCG AA - #T GAA AGA TGC CAG AGG           960                                                                        Leu Gln Glu Val Asp Val Glu Val Ile Pro As - #n Glu Arg Cys Gln Arg            305                 3 - #10                 3 - #15                 3 -       #20                                                                               - - TGG TTC CGT GCT GCG GGT CGA CGA GAA ACC AT - #T CAC GAT GTC TTT         CTC     1008                                                                     Trp Phe Arg Ala Ala Gly Arg Arg Glu Thr Il - #e His Asp Val Phe Leu                           325  - #               330  - #               335               - - TGC GCC GGA TAT AAA GAG GGT GGT CGT GAT TC - #A TGC CAA GGT GAT TCT          1056                                                                        Cys Ala Gly Tyr Lys Glu Gly Gly Arg Asp Se - #r Cys Gln Gly Asp Ser                        340      - #           345      - #           350                   - - GGA GGT CCT CTA ATA ATG CAG ATT GAG GGT AG - #A AGG ACC CTT GTG GGT          1104                                                                        Gly Gly Pro Leu Ile Met Gln Ile Glu Gly Ar - #g Arg Thr Leu Val Gly                    355          - #       360          - #       365                       - - CTA GTT TCT TGG GGC ATT GGA TGT GGT CGT GA - #G CAT TTA CCA GGC GTA          1152                                                                        Leu Val Ser Trp Gly Ile Gly Cys Gly Arg Gl - #u His Leu Pro Gly Val                370              - #   375              - #   380                           - - TAT ACC AAT ATA CAA AAA TTC ATA CCG TGG AT - #C GAC AAA GTA ATG GGA          1200                                                                        Tyr Thr Asn Ile Gln Lys Phe Ile Pro Trp Il - #e Asp Lys Val Met Gly            385                 3 - #90                 3 - #95                 4 -       #00                                                                               - -  - - (2) INFORMATION FOR SEQ ID NO:33:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 400 amino - #acids                                                 (B) TYPE: amino acid                                                           (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:33:                               - - Met Leu Ala Ile Val Pro Ser Asn Gly Ala Ph - #e Ala Asp His Ala         Asn                                                                                1               5 - #                 10 - #                 15              - - Leu Gly Gly Val Asp Gly Leu Ser Gly Leu Il - #e Leu Val Ala Val Ala                    20     - #             25     - #             30                   - - Ile Ser Ser Ile Gly Tyr Ala Asp Ala Ala As - #n Val Ala Gln Asp Gly                35         - #         40         - #         45                       - - His Pro Ser Ser Gln Gln Glu Gln Glu Ile Le - #u Leu Leu Asn Ala Leu            50             - #     55             - #     60                           - - Ala Arg Arg Asn Gly Ala Thr Gly His Gln Ph - #e Asp Val Asp Gln Asp        65                 - # 70                 - # 75                 - # 80        - - Ser Ile Met Asp Met Leu Gly Arg Met Ile Pr - #o Gln Thr Cys Arg Tyr                        85 - #                 90 - #                 95               - - Lys Gly Glu Arg Phe Glu Cys Gly Leu Ser Il - #e Ser Cys Val Leu Gly                   100      - #           105      - #           110                   - - Gly Gly Lys Pro Leu Asp Leu Cys Ser Gly Gl - #y Met Ile Trp Ser Cys               115          - #       120          - #       125                       - - Cys Val Asp Arg Asp Ile Arg Pro Glu Pro Gl - #n His Gln Gly Ala Leu           130              - #   135              - #   140                           - - Gln Asn Ala Thr Cys Gly Glu Leu Tyr Thr Ar - #g Ser Asn Arg Ile Val       145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - Gly Gly His Ser Thr Gly Phe Gly Ser His Pr - #o Trp Gln Ala Ala         Leu                                                                                              165  - #               170  - #               175              - - Ile Lys Ser Gly Phe Leu Ser Lys Lys Leu Se - #r Cys Gly Gly Ala Leu                   180      - #           185      - #           190                   - - Val Ser Asp Arg Trp Val Ile Thr Ala Ala Hi - #s Cys Val Ala Thr Thr               195          - #       200          - #       205                       - - Pro Asn Ser Asn Leu Lys Val Arg Leu Gly Gl - #u Trp Asp Val Arg Asp           210              - #   215              - #   220                           - - His Asp Glu Arg Leu Asn His Glu Glu Tyr Al - #a Ile Glu Arg Lys Glu       225                 2 - #30                 2 - #35                 2 -       #40                                                                               - - Val His Pro Ser Tyr Ser Pro Thr Asp Phe Ar - #g Asn Asp Val Ala         Leu                                                                                              245  - #               250  - #               255              - - Val Lys Leu Asp Arg Thr Val Ile Phe Lys Gl - #n His Ile Leu Pro Val                   260      - #           265      - #           270                   - - Cys Leu Pro His Lys Gln Met Lys Leu Ala Gl - #y Lys Met Ala Thr Val               275          - #       280          - #       285                       - - Ala Gly Trp Gly Arg Thr Arg His Gly Gln Se - #r Thr Val Pro Ala Val           290              - #   295              - #   300                           - - Leu Gln Glu Val Asp Val Glu Val Ile Pro As - #n Glu Arg Cys Gln Arg       305                 3 - #10                 3 - #15                 3 -       #20                                                                               - - Trp Phe Arg Ala Ala Gly Arg Arg Glu Thr Il - #e His Asp Val Phe         Leu                                                                                              325  - #               330  - #               335              - - Cys Ala Gly Tyr Lys Glu Gly Gly Arg Asp Se - #r Cys Gln Gly Asp Ser                   340      - #           345      - #           350                   - - Gly Gly Pro Leu Ile Met Gln Ile Glu Gly Ar - #g Arg Thr Leu Val Gly               355          - #       360          - #       365                       - - Leu Val Ser Trp Gly Ile Gly Cys Gly Arg Gl - #u His Leu Pro Gly Val           370              - #   375              - #   380                           - - Tyr Thr Asn Ile Gln Lys Phe Ile Pro Trp Il - #e Asp Lys Val Met Gly       385                 3 - #90                 3 - #95                 4 -       #00                                                                               - -  - - (2) INFORMATION FOR SEQ ID NO:34:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 1200 base - #pairs                                                 (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (genomic)                                      - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:34:                               - - TCCCATTACT TTGTCGATCC ACGGTATGAA TTTTTGTATA TTGGTATATA CG -             #CCTGGTAA     60                                                                  - - ATGCTCACGA CCACATCCAA TGCCCCAAGA AACTAGACCC ACAAGGGTCC TT -             #CTACCCTC    120                                                                  - - AATCTGCATT ATTAGAGGAC CTCCAGAATC ACCTTGGCAT GAATCACGAC CA -             #CCCTCTTT    180                                                                  - - ATATCCGGCG CAGAGAAAGA CATCGTGAAT GGTTTCTCGT CGACCCGCAG CA -             #CGGAACCA    240                                                                  - - CCTCTGGCAT CTTTCATTCG GAATCACCTC GACATCGACT TCTTGTAAGA CA -             #GCCGGCAC    300                                                                  - - AGTGCTCTGC CCGTGCCTCG TCCGTCCCCA TCCGGCGACT GTTGCCATTT TT -             #CCAGCCAG    360                                                                  - - TTTCATTTGC TTATGAGGTA AGCAGACAGG TAAAATATGT TGTTTGAAAA TA -             #ACAGTTCT    420                                                                  - - ATCGAGTTTC ACTAAGGCTA CATCATTCCG GAAATCGGTT GGTGAATATG AA -             #GGATGAAC    480                                                                  - - TTCTTTGCGT TCGATTGCGT ATTCCTCGTG GTTCAGTCGC TCATCGTGGT CG -             #CGGACGTC    540                                                                  - - CCATTCGCCC AATCGCACCT TCAGGTTCGA ATTTGGTGTC GTGGCAACGC AA -             #TGTGCAGC    600                                                                  - - AGTTATAACC CATCGATCGC TAACTAAAGC GCCACCGCAA GATAATTTTT TA -             #CTCAAAAA    660                                                                  - - TCCTGATTTG ATCAAAGCCG CCTGCCAAGG ATGAGACCCG AATCCTGTTG AA -             #TGACCTCC    720                                                                  - - TACGATTCTA TTAGACCTCG TGTACAATTC TCCACAAGTT GCGTTCTGCA GA -             #GCGCCCTG    780                                                                  - - GTGCTGCGGC TCAGGCCGAA TGTCCCTGTC GACGCAGCAC GACCAGATCA TT -             #CCGCCGCT    840                                                                  - - GCACAGGTCA AGAGGTTTTC CGCCGCCCAG GACGCACGAA ATTGACAAAC CG -             #CACTCGAA    900                                                                  - - CCGTTCGCCT TTGTACCGGC AAGTCTGAGG TATCATTCTT CCTAGCATAT CC -             #ATAATTGA    960                                                                  - - ATCTTGATCT ACGTCAAATT GGTGCCCCGT CGCTCCGTTC CTGCGAGCTA AG -             #GCATTCAG   1020                                                                  - - CAGCAGGATC TCCTGCTCTT GCTGGCTGGA CGGATGTCCG TCCTGCGCAA CG -             #TTCGCCGC   1080                                                                  - - GTCCGCATAT CCAATCGAAG ATATCGCAAC AGCGACCAGA ATCAAACCAG AA -             #AGACCATC   1140                                                                  - - AACTCCACCA AGGTTGGCAT GGTCTGCGAA CGCTCCGTTT GACGGGACGA TC -             #GCTAACAT   1200                                                                  - -  - - (2) INFORMATION FOR SEQ ID NO:35:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 726 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (genomic)                                      - -     (ix) FEATURE:                                                                   (A) NAME/KEY: CDS                                                              (B) LOCATION: 1..726                                                  - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:35:                               - - ATC GTA GGA GGT CAT TCA ACA GGA TTC GGG TC - #T CAT CCT TGG CAG         GCG       48                                                                     Ile Val Gly Gly His Ser Thr Gly Phe Gly Se - #r His Pro Trp Gln Ala             1               5 - #                 10 - #                 15               - - GCT TTG ATC AAA TCA GGA TTT TTG AGT AAA AA - #A TTA TCT TGC GGT GGC            96                                                                        Ala Leu Ile Lys Ser Gly Phe Leu Ser Lys Ly - #s Leu Ser Cys Gly Gly                         20     - #             25     - #             30                   - - GCT TTA GTT AGC GAT CGA TGG GTT ATA ACT GC - #T GCA CAT TGC GTT GCC           144                                                                        Ala Leu Val Ser Asp Arg Trp Val Ile Thr Al - #a Ala His Cys Val Ala                     35         - #         40         - #         45                       - - ACG ACA CCA AAT TCG AAC CTG AAG GTG CGA TT - #G GGC GAA TGG GAC GTC           192                                                                        Thr Thr Pro Asn Ser Asn Leu Lys Val Arg Le - #u Gly Glu Trp Asp Val                 50             - #     55             - #     60                           - - CGC GAC CAC GAT GAG CGA CTG AAC CAC GAG GA - #A TAC GCA ATC GAA CGC           240                                                                        Arg Asp His Asp Glu Arg Leu Asn His Glu Gl - #u Tyr Ala Ile Glu Arg             65                 - # 70                 - # 75                 - # 80        - - AAA GAA GTT CAT CCT TCA TAT TCA CCA ACC GA - #T TTC CGG AAT GAT GTA           288                                                                        Lys Glu Val His Pro Ser Tyr Ser Pro Thr As - #p Phe Arg Asn Asp Val                             85 - #                 90 - #                 95               - - GCC TTA GTG AAA CTC GAT AGA ACT GTT ATT TT - #C AAA CAA CAT ATT TTA           336                                                                        Ala Leu Val Lys Leu Asp Arg Thr Val Ile Ph - #e Lys Gln His Ile Leu                        100      - #           105      - #           110                   - - CCT GTC TGC TTA CCT CAT AAG CAA ATG AAA CT - #G GCT GGA AAA ATG GCA           384                                                                        Pro Val Cys Leu Pro His Lys Gln Met Lys Le - #u Ala Gly Lys Met Ala                    115          - #       120          - #       125                       - - ACA GTC GCC GGA TGG GGA CGG ACG AGG CAC GG - #G CAG AGC ACT GTG CCG           432                                                                        Thr Val Ala Gly Trp Gly Arg Thr Arg His Gl - #y Gln Ser Thr Val Pro                130              - #   135              - #   140                           - - GCT GTC TTA CAA GAA GTC GAT GTC GAG GTG AT - #T CCG AAT GAA AGA TGC           480                                                                        Ala Val Leu Gln Glu Val Asp Val Glu Val Il - #e Pro Asn Glu Arg Cys            145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - CAG AGG TGG TTC CGT GCT GCG GGT CGA CGA GA - #A ACC ATT CAC GAT         GTC      528                                                                     Gln Arg Trp Phe Arg Ala Ala Gly Arg Arg Gl - #u Thr Ile His Asp Val                           165  - #               170  - #               175               - - TTT CTC TGC GCC GGA TAT AAA GAG GGT GGT CG - #T GAT TCA TGC CAA GGT           576                                                                        Phe Leu Cys Ala Gly Tyr Lys Glu Gly Gly Ar - #g Asp Ser Cys Gln Gly                        180      - #           185      - #           190                   - - GAT TCT GGA GGT CCT CTA ATA ATG CAG ATT GA - #G GGT AGA AGG ACC CTT           624                                                                        Asp Ser Gly Gly Pro Leu Ile Met Gln Ile Gl - #u Gly Arg Arg Thr Leu                    195          - #       200          - #       205                       - - GTG GGT CTA GTT TCT TGG GGC ATT GGA TGT GG - #T CGT GAG CAT TTA CCA           672                                                                        Val Gly Leu Val Ser Trp Gly Ile Gly Cys Gl - #y Arg Glu His Leu Pro                210              - #   215              - #   220                           - - GGC GTA TAT ACC AAT ATA CAA AAA TTC ATA CC - #G TGG ATC GAC AAA GTA           720                                                                        Gly Val Tyr Thr Asn Ile Gln Lys Phe Ile Pr - #o Trp Ile Asp Lys Val            225                 2 - #30                 2 - #35                 2 -       #40                                                                               - - ATG GGA                - #                  - #                  -      #          726                                                                  Met Gly                                                                         - -  - - (2) INFORMATION FOR SEQ ID NO:36:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 242 amino - #acids                                                 (B) TYPE: amino acid                                                           (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:36:                               - - Ile Val Gly Gly His Ser Thr Gly Phe Gly Se - #r His Pro Trp Gln Ala         1               5 - #                 10 - #                 15               - - Ala Leu Ile Lys Ser Gly Phe Leu Ser Lys Ly - #s Leu Ser Cys Gly Gly                    20     - #             25     - #             30                   - - Ala Leu Val Ser Asp Arg Trp Val Ile Thr Al - #a Ala His Cys Val Ala                35         - #         40         - #         45                       - - Thr Thr Pro Asn Ser Asn Leu Lys Val Arg Le - #u Gly Glu Trp Asp Val            50             - #     55             - #     60                           - - Arg Asp His Asp Glu Arg Leu Asn His Glu Gl - #u Tyr Ala Ile Glu Arg        65                 - # 70                 - # 75                 - # 80        - - Lys Glu Val His Pro Ser Tyr Ser Pro Thr As - #p Phe Arg Asn Asp Val                        85 - #                 90 - #                 95               - - Ala Leu Val Lys Leu Asp Arg Thr Val Ile Ph - #e Lys Gln His Ile Leu                   100      - #           105      - #           110                   - - Pro Val Cys Leu Pro His Lys Gln Met Lys Le - #u Ala Gly Lys Met Ala               115          - #       120          - #       125                       - - Thr Val Ala Gly Trp Gly Arg Thr Arg His Gl - #y Gln Ser Thr Val Pro           130              - #   135              - #   140                           - - Ala Val Leu Gln Glu Val Asp Val Glu Val Il - #e Pro Asn Glu Arg Cys       145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - Gln Arg Trp Phe Arg Ala Ala Gly Arg Arg Gl - #u Thr Ile His Asp         Val                                                                                              165  - #               170  - #               175              - - Phe Leu Cys Ala Gly Tyr Lys Glu Gly Gly Ar - #g Asp Ser Cys Gln Gly                   180      - #           185      - #           190                   - - Asp Ser Gly Gly Pro Leu Ile Met Gln Ile Gl - #u Gly Arg Arg Thr Leu               195          - #       200          - #       205                       - - Val Gly Leu Val Ser Trp Gly Ile Gly Cys Gl - #y Arg Glu His Leu Pro           210              - #   215              - #   220                           - - Gly Val Tyr Thr Asn Ile Gln Lys Phe Ile Pr - #o Trp Ile Asp Lys Val       225                 2 - #30                 2 - #35                 2 -       #40                                                                               - - Met Gly                                                                    - -  - - (2) INFORMATION FOR SEQ ID NO:37:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 428 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: cDNA                                               - -     (ix) FEATURE:                                                                   (A) NAME/KEY: CDS                                                              (B) LOCATION: 1..428                                                  - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #any amino acid                                          (B) LOCATION: 17                                                      - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:37:                               - - GTA ACC AAA TAT GAA AAC GAT ATC GAA GGT TT - #A AAA GTT AGG GTT         GGA       48                                                                     Val Thr Lys Tyr Glu Asn Asp Ile Glu Gly Le - #u Lys Val Arg Val Gly             1               5 - #                 10 - #                 15               - - ANC AAT GAG CAT AAC AAA GGT GGG CGT TTA TA - #C GAC ATT AAA GAA ATT            96                                                                        Xaa Asn Glu His Asn Lys Gly Gly Arg Leu Ty - #r Asp Ile Lys Glu Ile                         20     - #             25     - #             30                   - - AAA AAA CAT CCA AGA TAT AAC GAT CGA ACC AG - #A TAC GAT TTT GAT GTC           144                                                                        Lys Lys His Pro Arg Tyr Asn Asp Arg Thr Ar - #g Tyr Asp Phe Asp Val                     35         - #         40         - #         45                       - - GCT TTA TTA CGC ATT GCA AAG CCA ATT GCA TA - #C ACT GCT TGC ACT GTT           192                                                                        Ala Leu Leu Arg Ile Ala Lys Pro Ile Ala Ty - #r Thr Ala Cys Thr Val                 50             - #     55             - #     60                           - - GTT CCT GTA GCA TTG GCA GAA ACT GGA AAA GA - #A GTT CCA GAA GGC GCA           240                                                                        Val Pro Val Ala Leu Ala Glu Thr Gly Lys Gl - #u Val Pro Glu Gly Ala             65                 - # 70                 - # 75                 - # 80        - - CTC GTT AGT GTC ACA GGA TGG GGG GCT ACT AT - #G GTG GGC GGC CCA GCA           288                                                                        Leu Val Ser Val Thr Gly Trp Gly Ala Thr Me - #t Val Gly Gly Pro Ala                             85 - #                 90 - #                 95               - - TCA ACG CAT CTA AAA GGT GTT AAG GTT CCA AT - #C GTG TCA AAT GAA GAA           336                                                                        Ser Thr His Leu Lys Gly Val Lys Val Pro Il - #e Val Ser Asn Glu Glu                        100      - #           105      - #           110                   - - TGC AAC AAA AAT TAT ACC ATT CCT GGA GGT CT - #G GAT GAC AAA ATT TCA           384                                                                        Cys Asn Lys Asn Tyr Thr Ile Pro Gly Gly Le - #u Asp Asp Lys Ile Ser                    115          - #       120          - #       125                       - - GAC AGC ATG TTT TGC GCT GGT TTC CCT GAA GG - #C GGA AAG GAC  TC               - #428                                                                     Asp Ser Met Phe Cys Ala Gly Phe Pro Glu Gl - #y Gly Lys Asp                        130              - #   135              - #   140                           - -  - - (2) INFORMATION FOR SEQ ID NO:38:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 142 amino - #acids                                                 (B) TYPE: amino acid                                                           (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #any amino acid                                          (B) LOCATION: 17                                                      - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:38:                               - - Val Thr Lys Tyr Glu Asn Asp Ile Glu Gly Le - #u Lys Val Arg Val Gly         1               5 - #                 10 - #                 15               - - Xaa Asn Glu His Asn Lys Gly Gly Arg Leu Ty - #r Asp Ile Lys Glu Ile                    20     - #             25     - #             30                   - - Lys Lys His Pro Arg Tyr Asn Asp Arg Thr Ar - #g Tyr Asp Phe Asp Val                35         - #         40         - #         45                       - - Ala Leu Leu Arg Ile Ala Lys Pro Ile Ala Ty - #r Thr Ala Cys Thr Val            50             - #     55             - #     60                           - - Val Pro Val Ala Leu Ala Glu Thr Gly Lys Gl - #u Val Pro Glu Gly Ala        65                 - # 70                 - # 75                 - # 80        - - Leu Val Ser Val Thr Gly Trp Gly Ala Thr Me - #t Val Gly Gly Pro Ala                        85 - #                 90 - #                 95               - - Ser Thr His Leu Lys Gly Val Lys Val Pro Il - #e Val Ser Asn Glu Glu                   100      - #           105      - #           110                   - - Cys Asn Lys Asn Tyr Thr Ile Pro Gly Gly Le - #u Asp Asp Lys Ile Ser               115          - #       120          - #       125                       - - Asp Ser Met Phe Cys Ala Gly Phe Pro Glu Gl - #y Gly Lys Asp                   130              - #   135              - #   140                           - -  - - (2) INFORMATION FOR SEQ ID NO:39:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 428 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (genomic)                                      - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:39:                               - - GAGTCCTTTC CGCCTTCAGG GAAACCAGCG CAAAACATGC TGTCTGAAAT TT -              #TGTCATCC     60                                                                  - - AGACCTCCAG GAATGGTATA ATTTTTGTTG CATTCTTCAT TTGACACGAT TG -             #GAACCTTA    120                                                                  - - ACACCTTTTA GATGCGTTGA TGCTGGGCCG CCCACCATAG TAGCCCCCCA TC -             #CTGTGACA    180                                                                  - - CTAACGAGTG CGCCTTCTGG AACTTCTTTT CCAGTTTCTG CCAATGCTAC AG -             #GAACAACA    240                                                                  - - GTGCAAGCAG TGTATGCAAT TGGCTTTGCA ATGCGTAATA AAGCGACATC AA -             #AATCGTAT    300                                                                  - - CTGGTTCGAT CGTTATATCT TGGATGTTTT TTAATTTCTT TAATGTCGTA TA -             #AACGCCCA    360                                                                  - - CCTTTGTTAT GCTCATTGNT TCCAACCCTA ACTTTTAAAC CTTCGATATC GT -             #TTTCATAT    420                                                                  - - TTGGTTAC                - #                  - #                        - #         428                                                                   - -  - - (2) INFORMATION FOR SEQ ID NO:40:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 841 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: cDNA                                               - -     (ix) FEATURE:                                                                   (A) NAME/KEY: CDS                                                              (B) LOCATION: 2..728                                                  - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #any amino acid                                          (B) LOCATION: 62                                                      - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #any amino acid                                          (B) LOCATION: 209                                                     - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:40:                               - - A GAT CAT CGA ATA GTA GGA GGT GAA GAT GTA - #GAT ATT TCA ACT TGT              46                                                                          Asp His Arg Ile Val Gly Gly Glu Asp V - #al Asp Ile Ser Thr Cys                  1              - # 5                 - # 10                 - # 15          - - GGA TGG CAA GTT TCG TTT CAC AAT AGG AAA GG - #A CAT TTT TGT GGA GGG            94                                                                        Gly Trp Gln Val Ser Phe His Asn Arg Lys Gl - #y His Phe Cys Gly Gly                             20 - #                 25 - #                 30               - - TCC ATC ATT GGC AAA GAA TGG ATT CTA ACT GC - #T GCG CAT TGT GTA ACC           142                                                                        Ser Ile Ile Gly Lys Glu Trp Ile Leu Thr Al - #a Ala His Cys Val Thr                         35     - #             40     - #             45                   - - AAA TAT GAA AAC GAT ATC GAA GGT TTA AAA GT - #T AGG GTT GGA ANC AAT           190                                                                        Lys Tyr Glu Asn Asp Ile Glu Gly Leu Lys Va - #l Arg Val Gly Xaa Asn                     50         - #         55         - #         60                       - - GAG CAT AAC AAA GGT GGG CGT TTA TAC GAC AT - #T AAA GAA ATT AAA AAA           238                                                                        Glu His Asn Lys Gly Gly Arg Leu Tyr Asp Il - #e Lys Glu Ile Lys Lys                 65             - #     70             - #     75                           - - CAT CCA AGA TAT AAC GAT CGA ACC AGA TAC GA - #T TTT GAT GTC GCT TTA           286                                                                        His Pro Arg Tyr Asn Asp Arg Thr Arg Tyr As - #p Phe Asp Val Ala Leu             80                 - # 85                 - # 90                 - # 95        - - TTA CGC ATT GCA AAG CCA ATT GCA TAC ACT GC - #T TGC ACT GTT GTT CCT           334                                                                        Leu Arg Ile Ala Lys Pro Ile Ala Tyr Thr Al - #a Cys Thr Val Val Pro                            100  - #               105  - #               110               - - GTA GCA TTG GCA GAA ACT GGA AAA GAA GTT CC - #A GAA GGC GCA CTC GTT           382                                                                        Val Ala Leu Ala Glu Thr Gly Lys Glu Val Pr - #o Glu Gly Ala Leu Val                        115      - #           120      - #           125                   - - AGT GTC ACA GGA TGG GGG GCT ACT ATG GTG GG - #C GGC CCA GCA TCA ACG           430                                                                        Ser Val Thr Gly Trp Gly Ala Thr Met Val Gl - #y Gly Pro Ala Ser Thr                    130          - #       135          - #       140                       - - CAT CTA AAA GGT GTT AAG GTT CCA ATC GTG TC - #A AAT GAA GAA TGC AAC           478                                                                        His Leu Lys Gly Val Lys Val Pro Ile Val Se - #r Asn Glu Glu Cys Asn                145              - #   150              - #   155                           - - AAA AAT TAT ACC ATT CCT GGA GGT CTG GAT GA - #C AAA ATT TCA GAC AGC           526                                                                        Lys Asn Tyr Thr Ile Pro Gly Gly Leu Asp As - #p Lys Ile Ser Asp Ser            160                 1 - #65                 1 - #70                 1 -       #75                                                                               - - ATG TTT TGC GCT GGT TTC CCT GAA GGC GGA AA - #G GAC TCG TGT CAA         GGA      574                                                                     Met Phe Cys Ala Gly Phe Pro Glu Gly Gly Ly - #s Asp Ser Cys Gln Gly                           180  - #               185  - #               190               - - GAT AGC GGT GGG CCT GTA GTG GAT GAA AAT AG - #G GTT CAG GTC GGA ATT           622                                                                        Asp Ser Gly Gly Pro Val Val Asp Glu Asn Ar - #g Val Gln Val Gly Ile                        195      - #           200      - #           205                   - - GTG TNT TGG GGC GAA GGC TGT GCT TTA GCA GG - #A AAA CCA GGC GTT TAT           670                                                                        Val Xaa Trp Gly Glu Gly Cys Ala Leu Ala Gl - #y Lys Pro Gly Val Tyr                    210          - #       215          - #       220                       - - GCA AAA GTT TCA CAT CCT GAC GTA AAA AGG TT - #T ATT GAA ACC GTA GCA           718                                                                        Ala Lys Val Ser His Pro Asp Val Lys Arg Ph - #e Ile Glu Thr Val Ala                225              - #   230              - #   235                           - - GGA ATC AAA T AAAATTTGTT AGAAAAAATG TAGACAAGTT GTAT - #AAACTN                 768                                                                        Gly Ile Lys                                                                    240                                                                             - - TCAATGAAAT TGTTTTATTT TTGGAAATAA AATATAATTT NTGAAAAAAA AA -              #AAAAAAAA    828                                                                  - - AAAAAAAAAA AAA              - #                  - #                       - #     841                                                                   - -  - - (2) INFORMATION FOR SEQ ID NO:41:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 242 amino - #acids                                                 (B) TYPE: amino acid                                                           (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #any amino acid                                          (B) LOCATION: 62                                                      - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #any amino acid                                          (B) LOCATION: 209                                                     - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:41:                               - - Asp His Arg Ile Val Gly Gly Glu Asp Val As - #p Ile Ser Thr Cys Gly         1               5 - #                 10 - #                 15               - - Trp Gln Val Ser Phe His Asn Arg Lys Gly Hi - #s Phe Cys Gly Gly Ser                    20     - #             25     - #             30                   - - Ile Ile Gly Lys Glu Trp Ile Leu Thr Ala Al - #a His Cys Val Thr Lys                35         - #         40         - #         45                       - - Tyr Glu Asn Asp Ile Glu Gly Leu Lys Val Ar - #g Val Gly Xaa Asn Glu            50             - #     55             - #     60                           - - His Asn Lys Gly Gly Arg Leu Tyr Asp Ile Ly - #s Glu Ile Lys Lys His        65                 - # 70                 - # 75                 - # 80        - - Pro Arg Tyr Asn Asp Arg Thr Arg Tyr Asp Ph - #e Asp Val Ala Leu Leu                        85 - #                 90 - #                 95               - - Arg Ile Ala Lys Pro Ile Ala Tyr Thr Ala Cy - #s Thr Val Val Pro Val                   100      - #           105      - #           110                   - - Ala Leu Ala Glu Thr Gly Lys Glu Val Pro Gl - #u Gly Ala Leu Val Ser               115          - #       120          - #       125                       - - Val Thr Gly Trp Gly Ala Thr Met Val Gly Gl - #y Pro Ala Ser Thr His           130              - #   135              - #   140                           - - Leu Lys Gly Val Lys Val Pro Ile Val Ser As - #n Glu Glu Cys Asn Lys       145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - Asn Tyr Thr Ile Pro Gly Gly Leu Asp Asp Ly - #s Ile Ser Asp Ser         Met                                                                                              165  - #               170  - #               175              - - Phe Cys Ala Gly Phe Pro Glu Gly Gly Lys As - #p Ser Cys Gln Gly Asp                   180      - #           185      - #           190                   - - Ser Gly Gly Pro Val Val Asp Glu Asn Arg Va - #l Gln Val Gly Ile Val               195          - #       200          - #       205                       - - Xaa Trp Gly Glu Gly Cys Ala Leu Ala Gly Ly - #s Pro Gly Val Tyr Ala           210              - #   215              - #   220                           - - Lys Val Ser His Pro Asp Val Lys Arg Phe Il - #e Glu Thr Val Ala Gly       225                 2 - #30                 2 - #35                 2 -       #40                                                                               - - Ile Lys                                                                    - -  - - (2) INFORMATION FOR SEQ ID NO:42:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 841 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (genomic)                                      - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:42:                               - - TTTTTTTTTT TTTTTTTTTT TTTTTTTTTT CANAAATTAT ATTTTATTTC CA -             #AAAATAAA     60                                                                  - - ACAATTTCAT TGANAGTTTA TACAACTTGT CTACATTTTT TCTAACAAAT TT -             #TATTTGAT    120                                                                  - - TCCTGCTACG GTTTCAATAA ACCTTTTTAC GTCAGGATGT GAAACTTTTG CA -             #TAAACGCC    180                                                                  - - TGGTTTTCCT GCTAAAGCAC AGCCTTCGCC CCAANACACA ATTCCGACCT GA -             #ACCCTATT    240                                                                  - - TTCATCCACT ACAGGCCCAC CGCTATCTCC TTGACACGAG TCCTTTCCGC CT -             #TCAGGGAA    300                                                                  - - ACCAGCGCAA AACATGCTGT CTGAAATTTT GTCATCCAGA CCTCCAGGAA TG -             #GTATAATT    360                                                                  - - TTTGTTGCAT TCTTCATTTG ACACGATTGG AACCTTAACA CCTTTTAGAT GC -             #GTTGATGC    420                                                                  - - TGGGCCGCCC ACCATAGTAG CCCCCCATCC TGTGACACTA ACGAGTGCGC CT -             #TCTGGAAC    480                                                                  - - TTCTTTTCCA GTTTCTGCCA ATGCTACAGG AACAACAGTG CAAGCAGTGT AT -             #GCAATTGG    540                                                                  - - CTTTGCAATG CGTAATAAAG CGACATCAAA ATCGTATCTG GTTCGATCGT TA -             #TATCTTGG    600                                                                  - - ATGTTTTTTA ATTTCTTTAA TGTCGTATAA ACGCCCACCT TTGTTATGCT CA -             #TTGNTTCC    660                                                                  - - AACCCTAACT TTTAAACCTT CGATATCGTT TTCATATTTG GTTACACAAT GC -             #GCAGCAGT    720                                                                  - - TAGAATCCAT TCTTTGCCAA TGATGGACCC TCCACAAAAA TGTCCTTTCC TA -             #TTGTGAAA    780                                                                  - - CGAAACTTGC CATCCACAAG TTGAAATATC TACATCTTCA CCTCCTACTA TT -             #CGATGATC    840                                                                  - - T                  - #                  - #                  - #                   841                                                                   - -  - - (2) INFORMATION FOR SEQ ID NO:43:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 717 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (genomic)                                      - -     (ix) FEATURE:                                                                   (A) NAME/KEY: CDS                                                              (B) LOCATION: 1..717                                                  - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #any amino acid                                          (B) LOCATION: 59                                                      - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #any amino acid                                          (B) LOCATION: 206                                                     - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:43:                               - - ATA GTA GGA GGT GAA GAT GTA GAT ATT TCA AC - #T TGT GGA TGG CAA GTT            48                                                                        Ile Val Gly Gly Glu Asp Val Asp Ile Ser Th - #r Cys Gly Trp Gln Val              1               5 - #                 10 - #                 15               - - TCG TTT CAC AAT AGG AAA GGA CAT TTT TGT GG - #A GGG TCC ATC ATT GGC            96                                                                        Ser Phe His Asn Arg Lys Gly His Phe Cys Gl - #y Gly Ser Ile Ile Gly                         20     - #             25     - #             30                   - - AAA GAA TGG ATT CTA ACT GCT GCG CAT TGT GT - #A ACC AAA TAT GAA AAC           144                                                                        Lys Glu Trp Ile Leu Thr Ala Ala His Cys Va - #l Thr Lys Tyr Glu Asn                     35         - #         40         - #         45                       - - GAT ATC GAA GGT TTA AAA GTT AGG GTT GGA AN - #C AAT GAG CAT AAC AAA           192                                                                        Asp Ile Glu Gly Leu Lys Val Arg Val Gly Xa - #a Asn Glu His Asn Lys                 50             - #     55             - #     60                           - - GGT GGG CGT TTA TAC GAC ATT AAA GAA ATT AA - #A AAA CAT CCA AGA TAT           240                                                                        Gly Gly Arg Leu Tyr Asp Ile Lys Glu Ile Ly - #s Lys His Pro Arg Tyr             65                 - # 70                 - # 75                 - # 80        - - AAC GAT CGA ACC AGA TAC GAT TTT GAT GTC GC - #T TTA TTA CGC ATT GCA           288                                                                        Asn Asp Arg Thr Arg Tyr Asp Phe Asp Val Al - #a Leu Leu Arg Ile Ala                             85 - #                 90 - #                 95               - - AAG CCA ATT GCA TAC ACT GCT TGC ACT GTT GT - #T CCT GTA GCA TTG GCA           336                                                                        Lys Pro Ile Ala Tyr Thr Ala Cys Thr Val Va - #l Pro Val Ala Leu Ala                        100      - #           105      - #           110                   - - GAA ACT GGA AAA GAA GTT CCA GAA GGC GCA CT - #C GTT AGT GTC ACA GGA           384                                                                        Glu Thr Gly Lys Glu Val Pro Glu Gly Ala Le - #u Val Ser Val Thr Gly                    115          - #       120          - #       125                       - - TGG GGG GCT ACT ATG GTG GGC GGC CCA GCA TC - #A ACG CAT CTA AAA GGT           432                                                                        Trp Gly Ala Thr Met Val Gly Gly Pro Ala Se - #r Thr His Leu Lys Gly                130              - #   135              - #   140                           - - GTT AAG GTT CCA ATC GTG TCA AAT GAA GAA TG - #C AAC AAA AAT TAT ACC           480                                                                        Val Lys Val Pro Ile Val Ser Asn Glu Glu Cy - #s Asn Lys Asn Tyr Thr            145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - ATT CCT GGA GGT CTG GAT GAC AAA ATT TCA GA - #C AGC ATG TTT TGC         GCT      528                                                                     Ile Pro Gly Gly Leu Asp Asp Lys Ile Ser As - #p Ser Met Phe Cys Ala                           165  - #               170  - #               175               - - GGT TTC CCT GAA GGC GGA AAG GAC TCG TGT CA - #A GGA GAT AGC GGT GGG           576                                                                        Gly Phe Pro Glu Gly Gly Lys Asp Ser Cys Gl - #n Gly Asp Ser Gly Gly                        180      - #           185      - #           190                   - - CCT GTA GTG GAT GAA AAT AGG GTT CAG GTC GG - #A ATT GTG TNT TGG GGC           624                                                                        Pro Val Val Asp Glu Asn Arg Val Gln Val Gl - #y Ile Val Xaa Trp Gly                    195          - #       200          - #       205                       - - GAA GGC TGT GCT TTA GCA GGA AAA CCA GGC GT - #T TAT GCA AAA GTT TCA           672                                                                        Glu Gly Cys Ala Leu Ala Gly Lys Pro Gly Va - #l Tyr Ala Lys Val Ser                210              - #   215              - #   220                           - - CAT CCT GAC GTA AAA AGG TTT ATT GAA ACC GT - #A GCA GGA ATC AAA               71 - #7                                                                    His Pro Asp Val Lys Arg Phe Ile Glu Thr Va - #l Ala Gly Ile Lys                225                 2 - #30                 2 - #35                             - -  - - (2) INFORMATION FOR SEQ ID NO:44:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 239 amino - #acids                                                 (B) TYPE: amino acid                                                           (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #any amino acid                                          (B) LOCATION: 59                                                      - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #any amino acid                                          (B) LOCATION: 206                                                     - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:44:                               - - Ile Val Gly Gly Glu Asp Val Asp Ile Ser Th - #r Cys Gly Trp Gln Val         1               5 - #                 10 - #                 15               - - Ser Phe His Asn Arg Lys Gly His Phe Cys Gl - #y Gly Ser Ile Ile Gly                    20     - #             25     - #             30                   - - Lys Glu Trp Ile Leu Thr Ala Ala His Cys Va - #l Thr Lys Tyr Glu Asn                35         - #         40         - #         45                       - - Asp Ile Glu Gly Leu Lys Val Arg Val Gly Xa - #a Asn Glu His Asn Lys            50             - #     55             - #     60                           - - Gly Gly Arg Leu Tyr Asp Ile Lys Glu Ile Ly - #s Lys His Pro Arg Tyr        65                 - # 70                 - # 75                 - # 80        - - Asn Asp Arg Thr Arg Tyr Asp Phe Asp Val Al - #a Leu Leu Arg Ile Ala                        85 - #                 90 - #                 95               - - Lys Pro Ile Ala Tyr Thr Ala Cys Thr Val Va - #l Pro Val Ala Leu Ala                   100      - #           105      - #           110                   - - Glu Thr Gly Lys Glu Val Pro Glu Gly Ala Le - #u Val Ser Val Thr Gly               115          - #       120          - #       125                       - - Trp Gly Ala Thr Met Val Gly Gly Pro Ala Se - #r Thr His Leu Lys Gly           130              - #   135              - #   140                           - - Val Lys Val Pro Ile Val Ser Asn Glu Glu Cy - #s Asn Lys Asn Tyr Thr       145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - Ile Pro Gly Gly Leu Asp Asp Lys Ile Ser As - #p Ser Met Phe Cys         Ala                                                                                              165  - #               170  - #               175              - - Gly Phe Pro Glu Gly Gly Lys Asp Ser Cys Gl - #n Gly Asp Ser Gly Gly                   180      - #           185      - #           190                   - - Pro Val Val Asp Glu Asn Arg Val Gln Val Gl - #y Ile Val Xaa Trp Gly               195          - #       200          - #       205                       - - Glu Gly Cys Ala Leu Ala Gly Lys Pro Gly Va - #l Tyr Ala Lys Val Ser           210              - #   215              - #   220                           - - His Pro Asp Val Lys Arg Phe Ile Glu Thr Va - #l Ala Gly Ile Lys           225                 2 - #30                 2 - #35                             - -  - - (2) INFORMATION FOR SEQ ID NO:45:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 717 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (genomic)                                      - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:45:                               - - TTTGATTCCT GCTACGGTTT CAATAAACCT TTTTACGTCA GGATGTGAAA CT -              #TTTGCATA     60                                                                  - - AACGCCTGGT TTTCCTGCTA AAGCACAGCC TTCGCCCCAA NACACAATTC CG -             #ACCTGAAC    120                                                                  - - CCTATTTTCA TCCACTACAG GCCCACCGCT ATCTCCTTGA CACGAGTCCT TT -             #CCGCCTTC    180                                                                  - - AGGGAAACCA GCGCAAAACA TGCTGTCTGA AATTTTGTCA TCCAGACCTC CA -             #GGAATGGT    240                                                                  - - ATAATTTTTG TTGCATTCTT CATTTGACAC GATTGGAACC TTAACACCTT TT -             #AGATGCGT    300                                                                  - - TGATGCTGGG CCGCCCACCA TAGTAGCCCC CCATCCTGTG ACACTAACGA GT -             #GCGCCTTC    360                                                                  - - TGGAACTTCT TTTCCAGTTT CTGCCAATGC TACAGGAACA ACAGTGCAAG CA -             #GTGTATGC    420                                                                  - - AATTGGCTTT GCAATGCGTA ATAAAGCGAC ATCAAAATCG TATCTGGTTC GA -             #TCGTTATA    480                                                                  - - TCTTGGATGT TTTTTAATTT CTTTAATGTC GTATAAACGC CCACCTTTGT TA -             #TGCTCATT    540                                                                  - - GNTTCCAACC CTAACTTTTA AACCTTCGAT ATCGTTTTCA TATTTGGTTA CA -             #CAATGCGC    600                                                                  - - AGCAGTTAGA ATCCATTCTT TGCCAATGAT GGACCCTCCA CAAAAATGTC CT -             #TTCCTATT    660                                                                  - - GTGAAACGAA ACTTGCCATC CACAAGTTGA AATATCTACA TCTTCACCTC CT - #ACTAT            717                                                                        - -  - - (2) INFORMATION FOR SEQ ID NO:46:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 20 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:46:                               - - GGACAAACTG TTCATTGCAG            - #                  - #                       - # 20                                                                    - -  - - (2) INFORMATION FOR SEQ ID NO:47:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 20 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:47:                               - - CCCTCATTTG TCGTAACTCC            - #                  - #                       - # 20                                                                    - -  - - (2) INFORMATION FOR SEQ ID NO:48:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 21 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:48:                               - - GGCTAGGTTA GTGGATTCTG G           - #                  - #                       - #21                                                                    - -  - - (2) INFORMATION FOR SEQ ID NO:49:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 27 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:49:                               - - GCAAATCAGT TCCAGAATCC ACTAACC          - #                  - #                  27                                                                       - -  - - (2) INFORMATION FOR SEQ ID NO:50:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 23 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:50:                               - - CAGGGCGCTC TGCAGAACGC AAC           - #                  - #                     23                                                                       - -  - - (2) INFORMATION FOR SEQ ID NO:51:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 22 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:51:                               - - ATTCCTCGTG GTTCAGTCGC TC           - #                  - #                      22                                                                       - -  - - (2) INFORMATION FOR SEQ ID NO:52:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 23 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:52:                               - - GGCAAGTTTC GTTTCACAAT AGG           - #                  - #                     23                                                                       - -  - - (2) INFORMATION FOR SEQ ID NO:53:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 24 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:53:                               - - TCCAACCCTA ACTTTTAAAC CTTC          - #                  - #                     24                                                                       - -  - - (2) INFORMATION FOR SEQ ID NO:54:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 31 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:54:                               - - CACAGGATCC AATAATTTGT GGTCAAAATG C        - #                  - #               31                                                                       - -  - - (2) INFORMATION FOR SEQ ID NO:55:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 37 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:55:                               - - AAAAAGAAAG CTTCTTTAAT TTTCTGACAT TGTCGTG      - #                        - #      37                                                                       - -  - - (2) INFORMATION FOR SEQ ID NO:56:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 31 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:56:                               - - GCGGGATCCT ATTGTGGGTG GTGAAGCAGT G        - #                  - #               31                                                                      - -  - - (2) INFORMATION FOR SEQ ID NO:57:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 35 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:57:                               - - GACGGTACCA TGTATAAAAT AATATTAAAC TCCGG       - #                  -      #       35                                                                       - -  - - (2) INFORMATION FOR SEQ ID NO:58:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 34 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:58:                               - - CCGGGATCCT ATGTTAGCGA TCGTCCCGTC AAAC       - #                  -       #        34                                                                       - -  - - (2) INFORMATION FOR SEQ ID NO:59:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 32 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:59:                               - - CCGGAATTCT TATCCCATTA CTTTGTCGAT CC       - #                  - #               32                                                                      - -  - - (2) INFORMATION FOR SEQ ID NO:60:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 32 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:60:                               - - GCGGGATCCA ATAGTAGGAG GTGAAGATGT AG       - #                  - #               32                                                                       - -  - - (2) INFORMATION FOR SEQ ID NO:61:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 36 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:61:                               - - CCGGAATTCT TCTAACAAAT TTTATTTGAT TCCTGC      - #                  -      #       36                                                                       - -  - - (2) INFORMATION FOR SEQ ID NO:62:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 26 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:62:                               - - GGATCCAATC GTTGGAGGTG AAGATG          - #                  - #                   26                                                                       - -  - - (2) INFORMATION FOR SEQ ID NO:63:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 27 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:63:                               - - GAATTCGAAA TCCACTTAAA CATTAGC          - #                  - #                  27                                                                       - -  - - (2) INFORMATION FOR SEQ ID NO:64:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 33 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:64:                               - - GATAAGGATC CGTTACCAGA TTCTTTCGAC TGG       - #                  - #              33                                                                       - -  - - (2) INFORMATION FOR SEQ ID NO:65:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 33 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:65:                               - - TTATCAAGCT TCCATTTACA TGCCGTAAAA ATC       - #                  - #              33                                                                       - -  - - (2) INFORMATION FOR SEQ ID NO:66:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 923 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: cDNA                                               - -     (ix) FEATURE:                                                                   (A) NAME/KEY: CDS                                                              (B) LOCATION: 2..802                                                  - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:66:                               - - A TTA ACA ATG ATG AAA CTT TTG GTA GTT TTT - #GCG ATT TTC GCT CAA              46                                                                           Leu Thr Met Met Lys Leu Leu Val Val P - #he Ala Ile Phe Ala Gln                  1              - # 5                 - # 10                 - # 15          - - ATC AGT TTT GTT TTT GGA AAT AAT GTA ACA GA - #A TTC GAT GAC CGA ATC            94                                                                        Ile Ser Phe Val Phe Gly Asn Asn Val Thr Gl - #u Phe Asp Asp Arg Ile                             20 - #                 25 - #                 30               - - GTT GGA GGT GAA GAT GTT GAT ATA TCA ACT TG - #T GGT TGG CAA ATT TCA           142                                                                        Val Gly Gly Glu Asp Val Asp Ile Ser Thr Cy - #s Gly Trp Gln Ile Ser                         35     - #             40     - #             45                   - - TTT CAA AGT GAA AAC CTT CAT TTT TGT GGA GG - #A TCA ATT ATT GCA CCA           190                                                                        Phe Gln Ser Glu Asn Leu His Phe Cys Gly Gl - #y Ser Ile Ile Ala Pro                     50         - #         55         - #         60                       - - AAA TGG ATT CTA ACT GCT GCA CAC TGT GTT GA - #A TGG TTG AAA AAG CCG           238                                                                        Lys Trp Ile Leu Thr Ala Ala His Cys Val Gl - #u Trp Leu Lys Lys Pro                 65             - #     70             - #     75                           - - CTC AAA GAC ATA ACC GTA CGT ATA GGA AGC AG - #T ATA CGT AAC AAA GGT           286                                                                        Leu Lys Asp Ile Thr Val Arg Ile Gly Ser Se - #r Ile Arg Asn Lys Gly             80                 - # 85                 - # 90                 - # 95        - - GGT CGA GTT CAT AAA GTA ATA GAT TTC CAC AT - #G CAT CCC TCG TAC AAT           334                                                                        Gly Arg Val His Lys Val Ile Asp Phe His Me - #t His Pro Ser Tyr Asn                            100  - #               105  - #               110               - - AAG AGG GCG GAT TAT GAT TTT GAC GTT GCT GT - #A CTA GAA CTT GAA AAA           382                                                                        Lys Arg Ala Asp Tyr Asp Phe Asp Val Ala Va - #l Leu Glu Leu Glu Lys                        115      - #           120      - #           125                   - - CCA GTC TCA TAT ACG GTT TGT ACA GTA GTA TC - #A GTA GAT TTA GCC GAA           430                                                                        Pro Val Ser Tyr Thr Val Cys Thr Val Val Se - #r Val Asp Leu Ala Glu                    130          - #       135          - #       140                       - - AGT GGA ACT GAA GTT AAA CCT GGA GCA ATA CT - #T AGT GTC ACT GGA TGG           478                                                                        Ser Gly Thr Glu Val Lys Pro Gly Ala Ile Le - #u Ser Val Thr Gly Trp                145              - #   150              - #   155                           - - GGT GCA ACT AAG GAA GGT GGT GGC GGA ACT TT - #G CAA CTA CAA GGT GTG           526                                                                        Gly Ala Thr Lys Glu Gly Gly Gly Gly Thr Le - #u Gln Leu Gln Gly Val            160                 1 - #65                 1 - #70                 1 -       #75                                                                               - - AAA GTT CCA GCT ATC TCT CCC AAA GAT TGT GC - #T AAG GGA TAT CCA         CCT      574                                                                     Lys Val Pro Ala Ile Ser Pro Lys Asp Cys Al - #a Lys Gly Tyr Pro Pro                           180  - #               185  - #               190               - - TCT GGA GGT AAA GAC AAA ATT ACA GAC AGC AT - #G TTA TGT GCT GGT CTT           622                                                                        Ser Gly Gly Lys Asp Lys Ile Thr Asp Ser Me - #t Leu Cys Ala Gly Leu                        195      - #           200      - #           205                   - - CCT GAA GGA GGT AAA GAT TCC TGC CAA GGC GA - #C AGT GGC GGT CCA CTG           670                                                                        Pro Glu Gly Gly Lys Asp Ser Cys Gln Gly As - #p Ser Gly Gly Pro Leu                    210          - #       215          - #       220                       - - GTA GAT GAA AAT AGA AAG CAA GTA GGA GTG GT - #T TCT TGG GGT CAA GGA           718                                                                        Val Asp Glu Asn Arg Lys Gln Val Gly Val Va - #l Ser Trp Gly Gln Gly                225              - #   230              - #   235                           - - TGT GCC AGA CCA GGA AAA CCA GGA ATT TAT GC - #T AAA GTG TCA CAC CCC           766                                                                        Cys Ala Arg Pro Gly Lys Pro Gly Ile Tyr Al - #a Lys Val Ser His Pro            240                 2 - #45                 2 - #50                 2 -       #55                                                                               - - GAA ATC AGA AAA TTT ATT GAA AAA TAT GCT AA - #T GTT TAAGTGGATT                812                                                                       Glu Ile Arg Lys Phe Ile Glu Lys Tyr Ala As - #n Val                                            260  - #               265                                      - - TCATTTTCAA TATAATGTGA TTTAAGATAC TCTTTAATGT TATGATATGA AT -              #TGTGATAA    872                                                                  - - ATTAAATAAT AAAGATTGAA GAAGTGATAA AAAAAAAAAA AAAAAAAAAA A - #                 923                                                                         - -  - - (2) INFORMATION FOR SEQ ID NO:67:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 267 amino - #acids                                                 (B) TYPE: amino acid                                                           (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:67:                               - - Leu Thr Met Met Lys Leu Leu Val Val Phe Al - #a Ile Phe Ala Gln Ile         1               5 - #                 10 - #                 15               - - Ser Phe Val Phe Gly Asn Asn Val Thr Glu Ph - #e Asp Asp Arg Ile Val                    20     - #             25     - #             30                   - - Gly Gly Glu Asp Val Asp Ile Ser Thr Cys Gl - #y Trp Gln Ile Ser Phe                35         - #         40         - #         45                       - - Gln Ser Glu Asn Leu His Phe Cys Gly Gly Se - #r Ile Ile Ala Pro Lys            50             - #     55             - #     60                           - - Trp Ile Leu Thr Ala Ala His Cys Val Glu Tr - #p Leu Lys Lys Pro Leu        65                 - # 70                 - # 75                 - # 80        - - Lys Asp Ile Thr Val Arg Ile Gly Ser Ser Il - #e Arg Asn Lys Gly Gly                        85 - #                 90 - #                 95               - - Arg Val His Lys Val Ile Asp Phe His Met Hi - #s Pro Ser Tyr Asn Lys                   100      - #           105      - #           110                   - - Arg Ala Asp Tyr Asp Phe Asp Val Ala Val Le - #u Glu Leu Glu Lys Pro               115          - #       120          - #       125                       - - Val Ser Tyr Thr Val Cys Thr Val Val Ser Va - #l Asp Leu Ala Glu Ser           130              - #   135              - #   140                           - - Gly Thr Glu Val Lys Pro Gly Ala Ile Leu Se - #r Val Thr Gly Trp Gly       145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - Ala Thr Lys Glu Gly Gly Gly Gly Thr Leu Gl - #n Leu Gln Gly Val         Lys                                                                                              165  - #               170  - #               175              - - Val Pro Ala Ile Ser Pro Lys Asp Cys Ala Ly - #s Gly Tyr Pro Pro Ser                   180      - #           185      - #           190                   - - Gly Gly Lys Asp Lys Ile Thr Asp Ser Met Le - #u Cys Ala Gly Leu Pro               195          - #       200          - #       205                       - - Glu Gly Gly Lys Asp Ser Cys Gln Gly Asp Se - #r Gly Gly Pro Leu Val           210              - #   215              - #   220                           - - Asp Glu Asn Arg Lys Gln Val Gly Val Val Se - #r Trp Gly Gln Gly Cys       225                 2 - #30                 2 - #35                 2 -       #40                                                                               - - Ala Arg Pro Gly Lys Pro Gly Ile Tyr Ala Ly - #s Val Ser His Pro         Glu                                                                                              245  - #               250  - #               255              - - Ile Arg Lys Phe Ile Glu Lys Tyr Ala Asn Va - #l                                       260      - #           265                                          - -  - - (2) INFORMATION FOR SEQ ID NO:68:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 15 amino - #acids                                                  (B) TYPE: amino acid                                                           (C) STRANDEDNESS:                                                              (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:68:                               - - Ile Val Gly Gly Glu Asp Val Asp Ile Ser Th - #r Cys Gly Trp Cys           1               5   - #                10  - #                15                - -  - - (2) INFORMATION FOR SEQ ID NO:69:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 34 amino - #acids                                                  (B) TYPE: amino acid                                                           (C) STRANDEDNESS:                                                              (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:69:                               - - Ile Val Gly Gly Glu Asp Val Asp Ile Ser Th - #r Cys Gly Trp Gln Ile       1               5   - #                10  - #                15                - - Ser Phe Gln Ser Glu Asn Leu His Phe Cys Gl - #y Gly Ser Ile Ile Ala                   20      - #            25      - #            30                    - - Pro Lys                                                                    - -  - - (2) INFORMATION FOR SEQ ID NO:70:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 15 amino - #acids                                                  (B) TYPE: amino acid                                                           (C) STRANDEDNESS:                                                              (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #His or Arg                                              (B) LOCATION: 1                                                       - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #Val or Pro                                              (B) LOCATION: 2                                                       - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #Gly or Ala or Ser                                       (B) LOCATION: 3                                                       - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #Tyr or Gly                                              (B) LOCATION: 4                                                       - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #Glu or Asn                                              (B) LOCATION: 5                                                       - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #Asp or Lys                                              (B) LOCATION: 6                                                       - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #Val or Arg                                              (B) LOCATION: 7                                                       - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #Asp or Ala                                              (B) LOCATION: 8                                                       - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #Asp or Pro                                              (B) LOCATION: 13                                                      - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:70:                               - - Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Asp Tyr As - #p Phe Xaa Val Ala           1               5   - #                10  - #                15                - -  - - (2) INFORMATION FOR SEQ ID NO:71:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 12 amino - #acids                                                  (B) TYPE: amino acid                                                           (C) STRANDEDNESS:                                                              (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #Ile or Gln                                              (B) LOCATION: 1                                                       - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #Tyr or Gly                                              (B) LOCATION: 4                                                       - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #Glu or Asn or Thr                                       (B) LOCATION: 5                                                       - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #Asp or Met or Pro                                       (B) LOCATION: 6                                                       - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #Lys or Asp                                              (B) LOCATION: 8                                                       - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #Asn or Ser                                              (B) LOCATION: 10                                                      - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #Met or Thr or Asn                                       (B) LOCATION: 11                                                      - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #Phe or Cys                                              (B) LOCATION: 12                                                      - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:71:                               - - Xaa Val Gly Xaa Xaa Xaa Val Xaa Ile Xaa Xa - #a Xaa                       1               5   - #                10                                       - -  - - (2) INFORMATION FOR SEQ ID NO:72:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 15 amino - #acids                                                  (B) TYPE: amino acid                                                           (C) STRANDEDNESS:                                                              (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #His or Arg                                              (B) LOCATION: 1                                                       - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #Ala or Ser                                              (B) LOCATION: 3                                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:72:                               - - Xaa Pro Xaa Tyr Asn Lys Arg Ala Asp Tyr As - #p Phe Asp Val Ala           1               5   - #                10  - #                15                - -  - - (2) INFORMATION FOR SEQ ID NO:73:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 28 amino - #acids                                                  (B) TYPE: amino acid                                                           (C) STRANDEDNESS:                                                              (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (ix) FEATURE:                                                                   (A) NAME/KEY: Xaa = - #probably Cys                                            (B) LOCATION: 1                                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:73:                               - - Xaa Pro Pro Pro Glu Met Leu Gly Gly Pro Se - #r Ile Phe Ile Phe Pro       1               5   - #                10  - #                15                - - Pro Lys Pro Lys Asp Asp Leu Leu Ile Lys Ar - #g Lys                                   20      - #            25                                           - -  - - (2) INFORMATION FOR SEQ ID NO:74:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 27 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:74:                               - - CTTTCCTCAC AATACCACCA AGGAAGC          - #                  - #                  27                                                                       - -  - - (2) INFORMATION FOR SEQ ID NO:75:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 23 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:75:                               - - CTTGTACGAT TGTCTCAACA GGC           - #                  - #                     23                                                                       - -  - - (2) INFORMATION FOR SEQ ID NO:76:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 573 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: cDNA                                               - -     (ix) FEATURE:                                                                   (A) NAME/KEY: CDS                                                              (B) LOCATION: 1..573                                                  - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:76:                               - - CGA TCT TTT AGA CAA GCG AAA TTG ATA ACG AA - #G TTT TCG AAG TCG GAT            48                                                                        Arg Ser Phe Arg Gln Ala Lys Leu Ile Thr Ly - #s Phe Ser Lys Ser Asp              1               5 - #                 10 - #                 15               - - GAA GTA AAA ACC TTG CGT TGG TTT CCC CGG TC - #C CAG GAT CAG GAA CAG            96                                                                        Glu Val Lys Thr Leu Arg Trp Phe Pro Arg Se - #r Gln Asp Gln Glu Gln                         20     - #             25     - #             30                   - - TTG CAC TTT ACC CCA ATG AGG GAA TTC GTG CA - #T CCC CAT TTT ACC GAA           144                                                                        Leu His Phe Thr Pro Met Arg Glu Phe Val Hi - #s Pro His Phe Thr Glu                     35         - #         40         - #         45                       - - CAT ATT GAT GAA GAA TTC CAC CGA TTC ATC AA - #T AAA CAC GGA AAA ATT           192                                                                        His Ile Asp Glu Glu Phe His Arg Phe Ile As - #n Lys His Gly Lys Ile                 50             - #     55             - #     60                           - - TAT AAT AAA AAT GAA GAA CAT CAT TTC CGC AA - #A GAA ATT TTC AGA CTA           240                                                                        Tyr Asn Lys Asn Glu Glu His His Phe Arg Ly - #s Glu Ile Phe Arg Leu             65                 - # 70                 - # 75                 - # 80        - - AAC TTG AGG TAC ATT TTT TCT AAG AAT CGT GC - #A AAT TTG GGA TAC ACT           288                                                                        Asn Leu Arg Tyr Ile Phe Ser Lys Asn Arg Al - #a Asn Leu Gly Tyr Thr                             85 - #                 90 - #                 95               - - TTG ACT GTT AAC CAT TTG GCT GAT CGT ACT GA - #A GCT GAA CTT AAG GCT           336                                                                        Leu Thr Val Asn His Leu Ala Asp Arg Thr Gl - #u Ala Glu Leu Lys Ala                        100      - #           105      - #           110                   - - TTG AGA GGA CAC AGA CCT TCC TCC GGT TAT AA - #T GGC GGT TTA CCC TTT           384                                                                        Leu Arg Gly His Arg Pro Ser Ser Gly Tyr As - #n Gly Gly Leu Pro Phe                    115          - #       120          - #       125                       - - CCT CAC AAT ACC ACC AAG GAA GCA AGA AAT TT - #A CCA GAT TCT TTC GAC           432                                                                        Pro His Asn Thr Thr Lys Glu Ala Arg Asn Le - #u Pro Asp Ser Phe Asp                130              - #   135              - #   140                           - - TGG CGA ATT TAT GGA GCT GTT ACT CCA GTT AA - #A GAT CAA TCT GTT TGT           480                                                                        Trp Arg Ile Tyr Gly Ala Val Thr Pro Val Ly - #s Asp Gln Ser Val Cys            145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - GGT TCC TGC TGG TCT TTC GGA ACA ATT GGA GC - #A ATC GAA GGT GCA         TAT      528                                                                     Gly Ser Cys Trp Ser Phe Gly Thr Ile Gly Al - #a Ile Glu Gly Ala Tyr                           165  - #               170  - #               175               - - TTC TTG AAA ACG GCG GTA ATC TGT ACG ATG TC - #T CAC AGC TTG ATG               57 - #3                                                                    Phe Leu Lys Thr Ala Val Ile Cys Thr Met Se - #r His Ser Leu Met                            180      - #           185      - #           190                   - -  - - (2) INFORMATION FOR SEQ ID NO:77:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 191 amino - #acids                                                 (B) TYPE: amino acid                                                           (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:77:                               - - Arg Ser Phe Arg Gln Ala Lys Leu Ile Thr Ly - #s Phe Ser Lys Ser Asp         1               5 - #                 10 - #                 15               - - Glu Val Lys Thr Leu Arg Trp Phe Pro Arg Se - #r Gln Asp Gln Glu Gln                    20     - #             25     - #             30                   - - Leu His Phe Thr Pro Met Arg Glu Phe Val Hi - #s Pro His Phe Thr Glu                35         - #         40         - #         45                       - - His Ile Asp Glu Glu Phe His Arg Phe Ile As - #n Lys His Gly Lys Ile            50             - #     55             - #     60                           - - Tyr Asn Lys Asn Glu Glu His His Phe Arg Ly - #s Glu Ile Phe Arg Leu        65                 - # 70                 - # 75                 - # 80        - - Asn Leu Arg Tyr Ile Phe Ser Lys Asn Arg Al - #a Asn Leu Gly Tyr Thr                        85 - #                 90 - #                 95               - - Leu Thr Val Asn His Leu Ala Asp Arg Thr Gl - #u Ala Glu Leu Lys Ala                   100      - #           105      - #           110                   - - Leu Arg Gly His Arg Pro Ser Ser Gly Tyr As - #n Gly Gly Leu Pro Phe               115          - #       120          - #       125                       - - Pro His Asn Thr Thr Lys Glu Ala Arg Asn Le - #u Pro Asp Ser Phe Asp           130              - #   135              - #   140                           - - Trp Arg Ile Tyr Gly Ala Val Thr Pro Val Ly - #s Asp Gln Ser Val Cys       145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - Gly Ser Cys Trp Ser Phe Gly Thr Ile Gly Al - #a Ile Glu Gly Ala         Tyr                                                                                              165  - #               170  - #               175              - - Phe Leu Lys Thr Ala Val Ile Cys Thr Met Se - #r His Ser Leu Met                       180      - #           185      - #           190                   - -  - - (2) INFORMATION FOR SEQ ID NO:78:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 410 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: cDNA                                               - -     (ix) FEATURE:                                                                   (A) NAME/KEY: CDS                                                              (B) LOCATION: 1..408                                                  - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:78:                               - - TGG GTT GTT ACT GCT GCT CAT TGT TTG AGA GG - #C AAA GAC CAC CTC CTG            48                                                                        Trp Val Val Thr Ala Ala His Cys Leu Arg Gl - #y Lys Asp His Leu Leu              1               5 - #                 10 - #                 15               - - GAC AAA CTG TTC ATT GCA GTC GGC CTG ACA AA - #T TTA GGT GAA GGA GGC            96                                                                        Asp Lys Leu Phe Ile Ala Val Gly Leu Thr As - #n Leu Gly Glu Gly Gly                         20     - #             25     - #             30                   - - ACC GTG TAT CCT GTA GAA AAA GGC ATC ATG CA - #C GAA GAA TAT GAA CAT           144                                                                        Thr Val Tyr Pro Val Glu Lys Gly Ile Met Hi - #s Glu Glu Tyr Glu His                     35         - #         40         - #         45                       - - TAT GAC ATA GTC AAC GAT ATT GCA CTA ATC AA - #A GTC AAA TCT CCG ATA           192                                                                        Tyr Asp Ile Val Asn Asp Ile Ala Leu Ile Ly - #s Val Lys Ser Pro Ile                 50             - #     55             - #     60                           - - GAA TTC AAT GAA AAA GTA ACG ACT GTA AAA TT - #A GGT GAG GAT TAT GTT           240                                                                        Glu Phe Asn Glu Lys Val Thr Thr Val Lys Le - #u Gly Glu Asp Tyr Val             65                 - # 70                 - # 75                 - # 80        - - GGC GGA GAC GTC CAA CTT CGA TTG ACA GGA TG - #G GGA GTT ACG ACA AAT           288                                                                        Gly Gly Asp Val Gln Leu Arg Leu Thr Gly Tr - #p Gly Val Thr Thr Asn                             85 - #                 90 - #                 95               - - GAG GGA ATC GGA AGC CCG AGT CAA AAA TTA CA - #G GTC ATG ACA GCC AAA           336                                                                        Glu Gly Ile Gly Ser Pro Ser Gln Lys Leu Gl - #n Val Met Thr Ala Lys                        100      - #           105      - #           110                   - - TCA CTA ACT TAT GAG GAT TGC AAA AAC GCA AT - #T TAT AAA AAA GAC TTT           384                                                                        Ser Leu Thr Tyr Glu Asp Cys Lys Asn Ala Il - #e Tyr Lys Lys Asp Phe                    115          - #       120          - #       125                       - - CGA AAG CCA AAT TTG TGC ACA GGC TA    - #                  - #                  410                                                                      Arg Lys Pro Asn Leu Cys Thr Gly                                                    130              - #   135                                                  - -  - - (2) INFORMATION FOR SEQ ID NO:79:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 136 amino - #acids                                                 (B) TYPE: amino acid                                                           (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:79:                               - - Trp Val Val Thr Ala Ala His Cys Leu Arg Gl - #y Lys Asp His Leu Leu         1               5 - #                 10 - #                 15               - - Asp Lys Leu Phe Ile Ala Val Gly Leu Thr As - #n Leu Gly Glu Gly Gly                    20     - #             25     - #             30                   - - Thr Val Tyr Pro Val Glu Lys Gly Ile Met Hi - #s Glu Glu Tyr Glu His                35         - #         40         - #         45                       - - Tyr Asp Ile Val Asn Asp Ile Ala Leu Ile Ly - #s Val Lys Ser Pro Ile            50             - #     55             - #     60                           - - Glu Phe Asn Glu Lys Val Thr Thr Val Lys Le - #u Gly Glu Asp Tyr Val        65                 - # 70                 - # 75                 - # 80        - - Gly Gly Asp Val Gln Leu Arg Leu Thr Gly Tr - #p Gly Val Thr Thr Asn                        85 - #                 90 - #                 95               - - Glu Gly Ile Gly Ser Pro Ser Gln Lys Leu Gl - #n Val Met Thr Ala Lys                   100      - #           105      - #           110                   - - Ser Leu Thr Tyr Glu Asp Cys Lys Asn Ala Il - #e Tyr Lys Lys Asp Phe               115          - #       120          - #       125                       - - Arg Lys Pro Asn Leu Cys Thr Gly                                               130              - #   135                                                  - -  - - (2) INFORMATION FOR SEQ ID NO:80:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 433 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: cDNA                                               - -     (ix) FEATURE:                                                                   (A) NAME/KEY: CDS                                                              (B) LOCATION: 1..432                                                  - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:80:                               - - GTA ACT GCT GCA CAT TGC TTT TAT GGA ACG TT - #A TTT CCG ATT GGA TTC            48                                                                        Val Thr Ala Ala His Cys Phe Tyr Gly Thr Le - #u Phe Pro Ile Gly Phe              1               5 - #                 10 - #                 15               - - TCT GCG AGA GCC GGC AGC AGT ACT GTG AAT TC - #A GGA GGA ACT GTG CAT            96                                                                        Ser Ala Arg Ala Gly Ser Ser Thr Val Asn Se - #r Gly Gly Thr Val His                         20     - #             25     - #             30                   - - ACA ATT TTG TAT TGG TAT ATT CAT CCA AAT TA - #T GAT TCA CAA AGT ACA           144                                                                        Thr Ile Leu Tyr Trp Tyr Ile His Pro Asn Ty - #r Asp Ser Gln Ser Thr                     35         - #         40         - #         45                       - - GAC TTT GAT GTT TCT GTA GTT CGA CTA TTA TC - #T TCT TTA AAT TTG AAT           192                                                                        Asp Phe Asp Val Ser Val Val Arg Leu Leu Se - #r Ser Leu Asn Leu Asn                 50             - #     55             - #     60                           - - GGA GGT TCT ATT CGA CCG GCT AGG TTA GTG GA - #T TCT GGA ACT GAT TTG           240                                                                        Gly Gly Ser Ile Arg Pro Ala Arg Leu Val As - #p Ser Gly Thr Asp Leu             65                 - # 70                 - # 75                 - # 80        - - CCA GCC GGT GAG ATG GTT ACA GTA ACT GGA TG - #G GGA CGA CTT TCG GAA           288                                                                        Pro Ala Gly Glu Met Val Thr Val Thr Gly Tr - #p Gly Arg Leu Ser Glu                             85 - #                 90 - #                 95               - - AAT ACT TCT GTT CCC TCG CCA TCA ACT CTT CA - #A GGA GTT ACA GTA CCA           336                                                                        Asn Thr Ser Val Pro Ser Pro Ser Thr Leu Gl - #n Gly Val Thr Val Pro                        100      - #           105      - #           110                   - - GTT GTA AGT AAT TCG GAA TGT CAA CAA CAA TT - #G CAA AAT CAG ACA ATC           384                                                                        Val Val Ser Asn Ser Glu Cys Gln Gln Gln Le - #u Gln Asn Gln Thr Ile                    115          - #       120          - #       125                       - - ACT GAC AAT ATG TTT TGT GCT GGT GAA TTA GA - #A GGA GGA AAG GAC TCT           432                                                                        Thr Asp Asn Met Phe Cys Ala Gly Glu Leu Gl - #u Gly Gly Lys Asp Ser                130              - #   135              - #   140                           - - T                  - #                  - #                  - #                   433                                                                    - -  - - (2) INFORMATION FOR SEQ ID NO:81:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 144 amino - #acids                                                 (B) TYPE: amino acid                                                           (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:81:                               - - Val Thr Ala Ala His Cys Phe Tyr Gly Thr Le - #u Phe Pro Ile Gly Phe         1               5 - #                 10 - #                 15               - - Ser Ala Arg Ala Gly Ser Ser Thr Val Asn Se - #r Gly Gly Thr Val His                    20     - #             25     - #             30                   - - Thr Ile Leu Tyr Trp Tyr Ile His Pro Asn Ty - #r Asp Ser Gln Ser Thr                35         - #         40         - #         45                       - - Asp Phe Asp Val Ser Val Val Arg Leu Leu Se - #r Ser Leu Asn Leu Asn            50             - #     55             - #     60                           - - Gly Gly Ser Ile Arg Pro Ala Arg Leu Val As - #p Ser Gly Thr Asp Leu        65                 - # 70                 - # 75                 - # 80        - - Pro Ala Gly Glu Met Val Thr Val Thr Gly Tr - #p Gly Arg Leu Ser Glu                        85 - #                 90 - #                 95               - - Asn Thr Ser Val Pro Ser Pro Ser Thr Leu Gl - #n Gly Val Thr Val Pro                   100      - #           105      - #           110                   - - Val Val Ser Asn Ser Glu Cys Gln Gln Gln Le - #u Gln Asn Gln Thr Ile               115          - #       120          - #       125                       - - Thr Asp Asn Met Phe Cys Ala Gly Glu Leu Gl - #u Gly Gly Lys Asp Ser           130              - #   135              - #   140                           - -  - - (2) INFORMATION FOR SEQ ID NO:82:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 426 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: cDNA                                               - -     (ix) FEATURE:                                                                   (A) NAME/KEY: CDS                                                              (B) LOCATION: 1..426                                                  - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:82:                               - - GCC ACG ACA CCA AAT TCG AAC CTG AAG GTG CG - #T TTG GGC GAA TGG GAC            48                                                                        Ala Thr Thr Pro Asn Ser Asn Leu Lys Val Ar - #g Leu Gly Glu Trp Asp              1               5 - #                 10 - #                 15               - - GTT CGC GAC CAC GAT GAG CGA CTG AAC CAC GA - #G GAA TAC GCA ATC GAA            96                                                                        Val Arg Asp His Asp Glu Arg Leu Asn His Gl - #u Glu Tyr Ala Ile Glu                         20     - #             25     - #             30                   - - CGC AAA GAA GTT CAT CCT TCA TAT TCA CCA AC - #C GAT TTC CGG AAT GAT           144                                                                        Arg Lys Glu Val His Pro Ser Tyr Ser Pro Th - #r Asp Phe Arg Asn Asp                     35         - #         40         - #         45                       - - GTA GCC TTA GTG AAA CTC GAT AGA ACT GTT AT - #T TTC AAA CAA CAT ATT           192                                                                        Val Ala Leu Val Lys Leu Asp Arg Thr Val Il - #e Phe Lys Gln His Ile                 50             - #     55             - #     60                           - - TTA CCT GTC TGC TTA CCT CAT AAG CAA ATG AA - #A CTG GCT GGA AAA ATG           240                                                                        Leu Pro Val Cys Leu Pro His Lys Gln Met Ly - #s Leu Ala Gly Lys Met             65                 - # 70                 - # 75                 - # 80        - - GCA ACA GTC GCC GGA TGG GGA CGG ACG AGG CA - #C GGG CAG AGC ACT GTG           288                                                                        Ala Thr Val Ala Gly Trp Gly Arg Thr Arg Hi - #s Gly Gln Ser Thr Val                             85 - #                 90 - #                 95               - - CCG GCT GTC TTA CAA GAA GTC GAT GTC GAG GT - #G ATT CCG AAT GAA AGA           336                                                                        Pro Ala Val Leu Gln Glu Val Asp Val Glu Va - #l Ile Pro Asn Glu Arg                        100      - #           105      - #           110                   - - TGC CAG AGG TGG TTC CGT GCT GCG GGT CGA CG - #A GAA ACC ATT CAC GAT           384                                                                        Cys Gln Arg Trp Phe Arg Ala Ala Gly Arg Ar - #g Glu Thr Ile His Asp                    115          - #       120          - #       125                       - - GTC TTT CTC TGC GCC GGA TAT AAA GAG GGT GG - #T CGT GAT TCA                  - # 426                                                                     Val Phe Leu Cys Ala Gly Tyr Lys Glu Gly Gl - #y Arg Asp Ser                        130              - #   135              - #   140                           - -  - - (2) INFORMATION FOR SEQ ID NO:83:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 142 amino - #acids                                                 (B) TYPE: amino acid                                                           (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:83:                               - - Ala Thr Thr Pro Asn Ser Asn Leu Lys Val Ar - #g Leu Gly Glu Trp Asp         1               5 - #                 10 - #                 15               - - Val Arg Asp His Asp Glu Arg Leu Asn His Gl - #u Glu Tyr Ala Ile Glu                    20     - #             25     - #             30                   - - Arg Lys Glu Val His Pro Ser Tyr Ser Pro Th - #r Asp Phe Arg Asn Asp                35         - #         40         - #         45                       - - Val Ala Leu Val Lys Leu Asp Arg Thr Val Il - #e Phe Lys Gln His Ile            50             - #     55             - #     60                           - - Leu Pro Val Cys Leu Pro His Lys Gln Met Ly - #s Leu Ala Gly Lys Met        65                 - # 70                 - # 75                 - # 80        - - Ala Thr Val Ala Gly Trp Gly Arg Thr Arg Hi - #s Gly Gln Ser Thr Val                        85 - #                 90 - #                 95               - - Pro Ala Val Leu Gln Glu Val Asp Val Glu Va - #l Ile Pro Asn Glu Arg                   100      - #           105      - #           110                   - - Cys Gln Arg Trp Phe Arg Ala Ala Gly Arg Ar - #g Glu Thr Ile His Asp               115          - #       120          - #       125                       - - Val Phe Leu Cys Ala Gly Tyr Lys Glu Gly Gl - #y Arg Asp Ser                   130              - #   135              - #   140                           - -  - - (2) INFORMATION FOR SEQ ID NO:84:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 778 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: cDNA                                               - -     (ix) FEATURE:                                                                   (A) NAME/KEY: CDS                                                              (B) LOCATION: 2..778                                                  - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:84:                               - - T GAT ACC TCA GAA TTG CCG GTA CAA AGG CGA - #ACG GTT TCG AGT GCG              46                                                                           Asp Thr Ser Glu Leu Pro Val Gln Arg A - #rg Thr Val Ser Ser Ala                  1              - # 5                 - # 10                 - # 15          - - GTT TGT CAA TTT TCG TGC GTC CTG GGC GGC GG - #A AAA CCT CTT GAC CTG            94                                                                        Val Cys Gln Phe Ser Cys Val Leu Gly Gly Gl - #y Lys Pro Leu Asp Leu                             20 - #                 25 - #                 30               - - TGC AGC GGC GGA ATG ATC TGG TCG TGC TGC GT - #C GAC AGG GAC ATT CGG           142                                                                        Cys Ser Gly Gly Met Ile Trp Ser Cys Cys Va - #l Asp Arg Asp Ile Arg                         35     - #             40     - #             45                   - - CCT GAG CCG CAG CAC CAG GGC GCT CTG CAG AA - #C GCA ACT TGT GGA GAA           190                                                                        Pro Glu Pro Gln His Gln Gly Ala Leu Gln As - #n Ala Thr Cys Gly Glu                     50         - #         55         - #         60                       - - TTG TAC ACG AGG TCT AAT AGA ATC GTA GGA GG - #T CAT TCA ACA GGA TTC           238                                                                        Leu Tyr Thr Arg Ser Asn Arg Ile Val Gly Gl - #y His Ser Thr Gly Phe                 65             - #     70             - #     75                           - - GGG TCT CAT CCT TGG CAG GCG GCT TTG ATC AA - #A TCA GGA TTT TTG AGT           286                                                                        Gly Ser His Pro Trp Gln Ala Ala Leu Ile Ly - #s Ser Gly Phe Leu Ser             80                 - # 85                 - # 90                 - # 95        - - AAA AAA TTA TCT TGC GGT GGC GCT TTA GTT AG - #C GAT CGA TGG GTT ATA           334                                                                        Lys Lys Leu Ser Cys Gly Gly Ala Leu Val Se - #r Asp Arg Trp Val Ile                            100  - #               105  - #               110               - - ACT GCT GCA CAT TGC GTT GCC ACG ACA CCA AA - #T TCG AAC CTG AAG GTG           382                                                                        Thr Ala Ala His Cys Val Ala Thr Thr Pro As - #n Ser Asn Leu Lys Val                        115      - #           120      - #           125                   - - CGA TTG GGC GAA TGG GAC GTC CGC GAC CAC GA - #T GAG CGA CTG AAC CAC           430                                                                        Arg Leu Gly Glu Trp Asp Val Arg Asp His As - #p Glu Arg Leu Asn His                    130          - #       135          - #       140                       - - GAG GAA TAC GCA ATC GAA CGC AAA GAA GTT CA - #T CCT TCA TAT TCA CCA           478                                                                        Glu Glu Tyr Ala Ile Glu Arg Lys Glu Val Hi - #s Pro Ser Tyr Ser Pro                145              - #   150              - #   155                           - - ACC GAT TTC CGG AAT GAT GTA GCC TTA GTG AA - #A CTC GAT AGA ACT GTT           526                                                                        Thr Asp Phe Arg Asn Asp Val Ala Leu Val Ly - #s Leu Asp Arg Thr Val            160                 1 - #65                 1 - #70                 1 -       #75                                                                               - - ATT TTC AAA CAA CAT ATT TTA CCT GTC TGC TT - #A CCT CAT AAG CAA         ATG      574                                                                     Ile Phe Lys Gln His Ile Leu Pro Val Cys Le - #u Pro His Lys Gln Met                           180  - #               185  - #               190               - - AAA CTG GCT GGA AAA ATG GCA ACA GTC GCC GG - #A TGG GGA CGG ACG AGG           622                                                                        Lys Leu Ala Gly Lys Met Ala Thr Val Ala Gl - #y Trp Gly Arg Thr Arg                        195      - #           200      - #           205                   - - CAC GGG CAG AGC ACT GTG CCG GCT GTC TTA CA - #A GAA GTC GAT GTC GAG           670                                                                        His Gly Gln Ser Thr Val Pro Ala Val Leu Gl - #n Glu Val Asp Val Glu                    210          - #       215          - #       220                       - - GTG ATT CCG AAT GAA AGA TGC CAG AGG TGG TT - #C CGT GCT GCG GGT CGA           718                                                                        Val Ile Pro Asn Glu Arg Cys Gln Arg Trp Ph - #e Arg Ala Ala Gly Arg                225              - #   230              - #   235                           - - CGA GAA ACC ATT CAC GAT GTC TTT CTC TGC GC - #C GGA TAT AAA GAG GGT           766                                                                        Arg Glu Thr Ile His Asp Val Phe Leu Cys Al - #a Gly Tyr Lys Glu Gly            240                 2 - #45                 2 - #50                 2 -       #55                                                                               - - GGT CGT GAT TCA            - #                  - #                       - #      778                                                                  Gly Arg Asp Ser                                                                 - -  - - (2) INFORMATION FOR SEQ ID NO:85:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 259 amino - #acids                                                 (B) TYPE: amino acid                                                           (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:85:                               - - Asp Thr Ser Glu Leu Pro Val Gln Arg Arg Th - #r Val Ser Ser Ala Val         1               5 - #                 10 - #                 15               - - Cys Gln Phe Ser Cys Val Leu Gly Gly Gly Ly - #s Pro Leu Asp Leu Cys                    20     - #             25     - #             30                   - - Ser Gly Gly Met Ile Trp Ser Cys Cys Val As - #p Arg Asp Ile Arg Pro                35         - #         40         - #         45                       - - Glu Pro Gln His Gln Gly Ala Leu Gln Asn Al - #a Thr Cys Gly Glu Leu            50             - #     55             - #     60                           - - Tyr Thr Arg Ser Asn Arg Ile Val Gly Gly Hi - #s Ser Thr Gly Phe Gly        65                 - # 70                 - # 75                 - # 80        - - Ser His Pro Trp Gln Ala Ala Leu Ile Lys Se - #r Gly Phe Leu Ser Lys                        85 - #                 90 - #                 95               - - Lys Leu Ser Cys Gly Gly Ala Leu Val Ser As - #p Arg Trp Val Ile Thr                   100      - #           105      - #           110                   - - Ala Ala His Cys Val Ala Thr Thr Pro Asn Se - #r Asn Leu Lys Val Arg               115          - #       120          - #       125                       - - Leu Gly Glu Trp Asp Val Arg Asp His Asp Gl - #u Arg Leu Asn His Glu           130              - #   135              - #   140                           - - Glu Tyr Ala Ile Glu Arg Lys Glu Val His Pr - #o Ser Tyr Ser Pro Thr       145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - Asp Phe Arg Asn Asp Val Ala Leu Val Lys Le - #u Asp Arg Thr Val         Ile                                                                                              165  - #               170  - #               175              - - Phe Lys Gln His Ile Leu Pro Val Cys Leu Pr - #o His Lys Gln Met Lys                   180      - #           185      - #           190                   - - Leu Ala Gly Lys Met Ala Thr Val Ala Gly Tr - #p Gly Arg Thr Arg His               195          - #       200          - #       205                       - - Gly Gln Ser Thr Val Pro Ala Val Leu Gln Gl - #u Val Asp Val Glu Val           210              - #   215              - #   220                           - - Ile Pro Asn Glu Arg Cys Gln Arg Trp Phe Ar - #g Ala Ala Gly Arg Arg       225                 2 - #30                 2 - #35                 2 -       #40                                                                               - - Glu Thr Ile His Asp Val Phe Leu Cys Ala Gl - #y Tyr Lys Glu Gly         Gly                                                                                              245  - #               250  - #               255              - - Arg Asp Ser                                                                - -  - - (2) INFORMATION FOR SEQ ID NO:86:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 24 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:86:                               - - TAAWGGWCCA GARTCTCCTT GACA          - #                  - #                     24                                                                       - -  - - (2) INFORMATION FOR SEQ ID NO:87:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 19 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:87:                               - - GGAAACAGCT ATGACCATG             - #                  - #                       - # 19                                                                    - -  - - (2) INFORMATION FOR SEQ ID NO:88:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 811 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (genomic)                                      - -     (ix) FEATURE:                                                                   (A) NAME/KEY: CDS                                                              (B) LOCATION: 1..762                                                  - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:88:                               - - ATG CGG GGT TCT CAT CAT CAT CAT CAT CAT GG - #T ATG GCT AGC ATG ACT            48                                                                        Met Arg Gly Ser His His His His His His Gl - #y Met Ala Ser Met Thr              1               5 - #                 10 - #                 15               - - GGT GGA CAG CAA ATG GGT CGG GAT CTG TAC GA - #C GAT GAC GAT AAG AAG            96                                                                        Gly Gly Gln Gln Met Gly Arg Asp Leu Tyr As - #p Asp Asp Asp Lys Lys                         20     - #             25     - #             30                   - - GAT CCG TTA CCA GAT TCT TTC GAC TGG AGA AT - #T TAT GGA GCT GTT ACT           144                                                                        Asp Pro Leu Pro Asp Ser Phe Asp Trp Arg Il - #e Tyr Gly Ala Val Thr                     35         - #         40         - #         45                       - - CCA GTT AAA GAT CAA TCT GTT TGT GGT TCC TG - #C TGG TCT TTC GGA ACA           192                                                                        Pro Val Lys Asp Gln Ser Val Cys Gly Ser Cy - #s Trp Ser Phe Gly Thr                 50             - #     55             - #     60                           - - ATT GGA GCT ATC GAA GGT GCA TAT TTC TTG AA - #A AAC GGC GGT AAT CTT           240                                                                        Ile Gly Ala Ile Glu Gly Ala Tyr Phe Leu Ly - #s Asn Gly Gly Asn Leu             65                 - # 70                 - # 75                 - # 80        - - GTA CGA TTG TCT CAA CAG GCT TTG ATT GAT TG - #T TCT TGG GGA TAT GGA           288                                                                        Val Arg Leu Ser Gln Gln Ala Leu Ile Asp Cy - #s Ser Trp Gly Tyr Gly                             85 - #                 90 - #                 95               - - AAT AAT GGT TGT GAC GGT GGC GAG GAC TTC CG - #C GCT TAC CAA TGG ATG           336                                                                        Asn Asn Gly Cys Asp Gly Gly Glu Asp Phe Ar - #g Ala Tyr Gln Trp Met                        100      - #           105      - #           110                   - - ATG AAA CAT GGA GGA ATT CCT ACT GAA GAA GA - #T TAT GGT GGT TAC TTG           384                                                                        Met Lys His Gly Gly Ile Pro Thr Glu Glu As - #p Tyr Gly Gly Tyr Leu                    115          - #       120          - #       125                       - - GGA CAA GAT GGT TAC TGC CAT GTC AAC AAC GT - #T ACT TTA GTT GCT CCC           432                                                                        Gly Gln Asp Gly Tyr Cys His Val Asn Asn Va - #l Thr Leu Val Ala Pro                130              - #   135              - #   140                           - - ATC ACA GGG TAT GTC AAC GTA ACA CGT AAT GA - #C GTT GAC GCT ATG AAG           480                                                                        Ile Thr Gly Tyr Val Asn Val Thr Arg Asn As - #p Val Asp Ala Met Lys            145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - GTT GCC CTT CTT AAA CAC GGT CCA ATT TCG GT - #G GCC ATT GAC GCA         TCA      528                                                                     Val Ala Leu Leu Lys His Gly Pro Ile Ser Va - #l Ala Ile Asp Ala Ser                           165  - #               170  - #               175               - - CAC AAA ACA TTC AGT TTT TAC TCC AAC GGC GT - #T TAC TAC GAA CCG AAA           576                                                                        His Lys Thr Phe Ser Phe Tyr Ser Asn Gly Va - #l Tyr Tyr Glu Pro Lys                        180      - #           185      - #           190                   - - TGT GGC AAT AAA GAG GAC GAG TTG GAC CAT GC - #C GTA TTA GTA GTC GGT           624                                                                        Cys Gly Asn Lys Glu Asp Glu Leu Asp His Al - #a Val Leu Val Val Gly                    195          - #       200          - #       205                       - - TAT GGT GAA ATC AAC AAC GAA CCT TAC TGG TT - #G GTC AAG AAT TCC TGG           672                                                                        Tyr Gly Glu Ile Asn Asn Glu Pro Tyr Trp Le - #u Val Lys Asn Ser Trp                210              - #   215              - #   220                           - - TCG AAT TTG TGG GGA AAT GAT GGT TAT ATT TT - #G ATG TCC GCC AGA AAT           720                                                                        Ser Asn Leu Trp Gly Asn Asp Gly Tyr Ile Le - #u Met Ser Ala Arg Asn            225                 2 - #30                 2 - #35                 2 -       #40                                                                               - - AAT AAT TGC GGA GTT TTG ACT GAT CCA ACT TA - #T GTT ACT ATG                  - # 762                                                                    Asn Asn Cys Gly Val Leu Thr Asp Pro Thr Ty - #r Val Thr Met                                    245  - #               250                                      - - TAACTTACTT TTTAGGAATT TGATTTTTAC GGCATGTAAA TGGAAGCTT  - #                   811                                                                          - -  - - (2) INFORMATION FOR SEQ ID NO:89:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 254 amino - #acids                                                 (B) TYPE: amino acid                                                           (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:89:                               - - Met Arg Gly Ser His His His His His His Gl - #y Met Ala Ser Met Thr         1               5 - #                 10 - #                 15               - - Gly Gly Gln Gln Met Gly Arg Asp Leu Tyr As - #p Asp Asp Asp Lys Lys                    20     - #             25     - #             30                   - - Asp Pro Leu Pro Asp Ser Phe Asp Trp Arg Il - #e Tyr Gly Ala Val Thr                35         - #         40         - #         45                       - - Pro Val Lys Asp Gln Ser Val Cys Gly Ser Cy - #s Trp Ser Phe Gly Thr            50             - #     55             - #     60                           - - Ile Gly Ala Ile Glu Gly Ala Tyr Phe Leu Ly - #s Asn Gly Gly Asn Leu        65                 - # 70                 - # 75                 - # 80        - - Val Arg Leu Ser Gln Gln Ala Leu Ile Asp Cy - #s Ser Trp Gly Tyr Gly                        85 - #                 90 - #                 95               - - Asn Asn Gly Cys Asp Gly Gly Glu Asp Phe Ar - #g Ala Tyr Gln Trp Met                   100      - #           105      - #           110                   - - Met Lys His Gly Gly Ile Pro Thr Glu Glu As - #p Tyr Gly Gly Tyr Leu               115          - #       120          - #       125                       - - Gly Gln Asp Gly Tyr Cys His Val Asn Asn Va - #l Thr Leu Val Ala Pro           130              - #   135              - #   140                           - - Ile Thr Gly Tyr Val Asn Val Thr Arg Asn As - #p Val Asp Ala Met Lys       145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - Val Ala Leu Leu Lys His Gly Pro Ile Ser Va - #l Ala Ile Asp Ala         Ser                                                                                              165  - #               170  - #               175              - - His Lys Thr Phe Ser Phe Tyr Ser Asn Gly Va - #l Tyr Tyr Glu Pro Lys                   180      - #           185      - #           190                   - - Cys Gly Asn Lys Glu Asp Glu Leu Asp His Al - #a Val Leu Val Val Gly               195          - #       200          - #       205                       - - Tyr Gly Glu Ile Asn Asn Glu Pro Tyr Trp Le - #u Val Lys Asn Ser Trp           210              - #   215              - #   220                           - - Ser Asn Leu Trp Gly Asn Asp Gly Tyr Ile Le - #u Met Ser Ala Arg Asn       225                 2 - #30                 2 - #35                 2 -       #40                                                                               - - Asn Asn Cys Gly Val Leu Thr Asp Pro Thr Ty - #r Val Thr Met                              245  - #               250                                      - -  - - (2) INFORMATION FOR SEQ ID NO:90:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 823 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (genomic)                                      - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:90:                               - - CCATTTACAT GCCGTAAAAA TCAAATTCCT AAAAAGTAAG TTACATAGTA AC -              #ATAAGTTG     60                                                                  - - GATCAGTCAA AACTCCGCAA TTATTATTTC TGGCGGACAT CAAAATATAA CC -             #ATCATTTC    120                                                                  - - CCCACAAATT CGACCAGGAA TTCTTGACCA ACCAGTAAGG TTCGTTGTTG AT -             #TTCACCAT    180                                                                  - - AACCGACTAC TAATACGGCA TGGTCCAACT CGTCCTCTTT ATTGCCACAT TT -             #CGGTTCGT    240                                                                  - - AGTAAACGCC GTTGGAGTAA AAACTGAATG TTTTGTGTGA TGCGTCAATG GC -             #CACCGAAA    300                                                                  - - TTGGACCGTG TTTAAGAAGG GCAACCTTCA TAGCGTCAAC GTCATTACGT GT -             #TACGTTGA    360                                                                  - - CATACCCTGT GATGGGAGCA ACTAAAGTAA CGTTGTTGAC ATGGCAGTAA CC -             #ATCTTGTC    420                                                                  - - CCAAGTAACC ACCATAATCT TCTTCAGTAG GAATTCCTCC ATGTTTCATC AT -             #CCATTGGT    480                                                                  - - AAGCGCGGAA GTCCTCGCCA CCGTCACAAC CATTATTTCC ATATCCCCAA GA -             #ACAATCAA    540                                                                  - - TCAAAGCCTG TTGAGACAAT CGTACAAGAT TACCGCCGTT TTTCAAGAAA TA -             #TGCACCTT    600                                                                  - - CGATAGCTCC AATTGTTCCG AAAGACCAGC AGGAACCACA AACAGATTGA TC -             #TTTAACTG    660                                                                  - - GAGTAACAGC TCCATAAATT CTCCAGTCGA AAGAATCTGG TAACGGATCC TT -             #ATCTGGTA    720                                                                  - - ACGGATCCTT CTTATCGTCA TCGTCGTACA GATCCCGACC CATTTCCTGT CC -             #ACCAGTCA    780                                                                  - - TGCTAGCCAT ACCATGATGA TGATGATGAT GAGAACCCCG CAT    - #                       - #823                                                                      - -  - - (2) INFORMATION FOR SEQ ID NO:91:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 762 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (genomic)                                      - -     (ix) FEATURE:                                                                   (A) NAME/KEY: CDS                                                              (B) LOCATION: 1..762                                                  - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:91:                               - - ATG CGG GGT TCT CAT CAT CAT CAT CAT CAT GG - #T ATG GCT AGC ATG ACT            48                                                                        Met Arg Gly Ser His His His His His His Gl - #y Met Ala Ser Met Thr              1               5 - #                 10 - #                 15               - - GGT GGA CAG CAA ATG GGT CGG GAT CTG TAC GA - #C GAT GAC GAT AAG AAG            96                                                                        Gly Gly Gln Gln Met Gly Arg Asp Leu Tyr As - #p Asp Asp Asp Lys Lys                         20     - #             25     - #             30                   - - GAT CCG TTA CCA GAT TCT TTC GAC TGG AGA AT - #T TAT GGA GCT GTT ACT           144                                                                        Asp Pro Leu Pro Asp Ser Phe Asp Trp Arg Il - #e Tyr Gly Ala Val Thr                     35         - #         40         - #         45                       - - CCA GTT AAA GAT CAA TCT GTT TGT GGT TCC TG - #C TGG TCT TTC GGA ACA           192                                                                        Pro Val Lys Asp Gln Ser Val Cys Gly Ser Cy - #s Trp Ser Phe Gly Thr                 50             - #     55             - #     60                           - - ATT GGA GCT ATC GAA GGT GCA TAT TTC TTG AA - #A AAC GGC GGT AAT CTT           240                                                                        Ile Gly Ala Ile Glu Gly Ala Tyr Phe Leu Ly - #s Asn Gly Gly Asn Leu             65                 - # 70                 - # 75                 - # 80        - - GTA CGA TTG TCT CAA CAG GCT TTG ATT GAT TG - #T TCT TGG GGA TAT GGA           288                                                                        Val Arg Leu Ser Gln Gln Ala Leu Ile Asp Cy - #s Ser Trp Gly Tyr Gly                             85 - #                 90 - #                 95               - - AAT AAT GGT TGT GAC GGT GGC GAG GAC TTC CG - #C GCT TAC CAA TGG ATG           336                                                                        Asn Asn Gly Cys Asp Gly Gly Glu Asp Phe Ar - #g Ala Tyr Gln Trp Met                        100      - #           105      - #           110                   - - ATG AAA CAT GGA GGA ATT CCT ACT GAA GAA GA - #T TAT GGT GGT TAC TTG           384                                                                        Met Lys His Gly Gly Ile Pro Thr Glu Glu As - #p Tyr Gly Gly Tyr Leu                    115          - #       120          - #       125                       - - GGA CAA GAT GGT TAC TGC CAT GTC AAC AAC GT - #T ACT TTA GTT GCT CCC           432                                                                        Gly Gln Asp Gly Tyr Cys His Val Asn Asn Va - #l Thr Leu Val Ala Pro                130              - #   135              - #   140                           - - ATC ACA GGG TAT GTC AAC GTA ACA CGT AAT GA - #C GTT GAC GCT ATG AAG           480                                                                        Ile Thr Gly Tyr Val Asn Val Thr Arg Asn As - #p Val Asp Ala Met Lys            145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - GTT GCC CTT CTT AAA CAC GGT CCA ATT TCG GT - #G GCC ATT GAC GCA         TCA      528                                                                     Val Ala Leu Leu Lys His Gly Pro Ile Ser Va - #l Ala Ile Asp Ala Ser                           165  - #               170  - #               175               - - CAC AAA ACA TTC AGT TTT TAC TCC AAC GGC GT - #T TAC TAC GAA CCG AAA           576                                                                        His Lys Thr Phe Ser Phe Tyr Ser Asn Gly Va - #l Tyr Tyr Glu Pro Lys                        180      - #           185      - #           190                   - - TGT GGC AAT AAA GAG GAC GAG TTG GAC CAT GC - #C GTA TTA GTA GTC GGT           624                                                                        Cys Gly Asn Lys Glu Asp Glu Leu Asp His Al - #a Val Leu Val Val Gly                    195          - #       200          - #       205                       - - TAT GGT GAA ATC AAC AAC GAA CCT TAC TGG TT - #G GTC AAG AAT TCC TGG           672                                                                        Tyr Gly Glu Ile Asn Asn Glu Pro Tyr Trp Le - #u Val Lys Asn Ser Trp                210              - #   215              - #   220                           - - TCG AAT TTG TGG GGA AAT GAT GGT TAT ATT TT - #G ATG TCC GCC AGA AAT           720                                                                        Ser Asn Leu Trp Gly Asn Asp Gly Tyr Ile Le - #u Met Ser Ala Arg Asn            225                 2 - #30                 2 - #35                 2 -       #40                                                                               - - AAT AAT TGC GGA GTT TTG ACT GAT CCA ACT TA - #T GTT ACT ATG                  - # 762                                                                    Asn Asn Cys Gly Val Leu Thr Asp Pro Thr Ty - #r Val Thr Met                                    245  - #               250                                      - -  - - (2) INFORMATION FOR SEQ ID NO:92:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 254 amino - #acids                                                 (B) TYPE: amino acid                                                           (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:92:                               - - Met Arg Gly Ser His His His His His His Gl - #y Met Ala Ser Met Thr         1               5 - #                 10 - #                 15               - - Gly Gly Gln Gln Met Gly Arg Asp Leu Tyr As - #p Asp Asp Asp Lys Lys                    20     - #             25     - #             30                   - - Asp Pro Leu Pro Asp Ser Phe Asp Trp Arg Il - #e Tyr Gly Ala Val Thr                35         - #         40         - #         45                       - - Pro Val Lys Asp Gln Ser Val Cys Gly Ser Cy - #s Trp Ser Phe Gly Thr            50             - #     55             - #     60                           - - Ile Gly Ala Ile Glu Gly Ala Tyr Phe Leu Ly - #s Asn Gly Gly Asn Leu        65                 - # 70                 - # 75                 - # 80        - - Val Arg Leu Ser Gln Gln Ala Leu Ile Asp Cy - #s Ser Trp Gly Tyr Gly                        85 - #                 90 - #                 95               - - Asn Asn Gly Cys Asp Gly Gly Glu Asp Phe Ar - #g Ala Tyr Gln Trp Met                   100      - #           105      - #           110                   - - Met Lys His Gly Gly Ile Pro Thr Glu Glu As - #p Tyr Gly Gly Tyr Leu               115          - #       120          - #       125                       - - Gly Gln Asp Gly Tyr Cys His Val Asn Asn Va - #l Thr Leu Val Ala Pro           130              - #   135              - #   140                           - - Ile Thr Gly Tyr Val Asn Val Thr Arg Asn As - #p Val Asp Ala Met Lys       145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - Val Ala Leu Leu Lys His Gly Pro Ile Ser Va - #l Ala Ile Asp Ala         Ser                                                                                              165  - #               170  - #               175              - - His Lys Thr Phe Ser Phe Tyr Ser Asn Gly Va - #l Tyr Tyr Glu Pro Lys                   180      - #           185      - #           190                   - - Cys Gly Asn Lys Glu Asp Glu Leu Asp His Al - #a Val Leu Val Val Gly               195          - #       200          - #       205                       - - Tyr Gly Glu Ile Asn Asn Glu Pro Tyr Trp Le - #u Val Lys Asn Ser Trp           210              - #   215              - #   220                           - - Ser Asn Leu Trp Gly Asn Asp Gly Tyr Ile Le - #u Met Ser Ala Arg Asn       225                 2 - #30                 2 - #35                 2 -       #40                                                                               - - Asn Asn Cys Gly Val Leu Thr Asp Pro Thr Ty - #r Val Thr Met                              245  - #               250                                      - -  - - (2) INFORMATION FOR SEQ ID NO:93:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 783 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (genomic)                                      - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:93:                               - - TTACATAGTA ACATAAGTTG GATCAGTCAA AACTCCGCAA TTATTATTTC TG -              #GCGGACAT     60                                                                  - - CAAAATATAA CCATCATTTC CCCACAAATT CGACCAGGAA TTCTTGACCA AC -             #CAGTAAGG    120                                                                  - - TTCGTTGTTG ATTTCACCAT AACCGACTAC TAATACGGCA TGGTCCAACT CG -             #TCCTCTTT    180                                                                  - - ATTGCCACAT TTCGGTTCGT AGTAAACGCC GTTGGAGTAA AAACTGAATG TT -             #TTGTGTGA    240                                                                  - - TGCGTCAATG GCCACCGAAA TTGGACCGTG TTTAAGAAGG GCAACCTTCA TA -             #GCGTCAAC    300                                                                  - - GTCATTACGT GTTACGTTGA CATACCCTGT GATGGGAGCA ACTAAAGTAA CG -             #TTGTTGAC    360                                                                  - - ATGGCAGTAA CCATCTTGTC CCAAGTAACC ACCATAATCT TCTTCAGTAG GA -             #ATTCCTCC    420                                                                  - - ATGTTTCATC ATCCATTGGT AAGCGCGGAA GTCCTCGCCA CCGTCACAAC CA -             #TTATTTCC    480                                                                  - - ATATCCCCAA GAACAATCAA TCAAAGCCTG TTGAGACAAT CGTACAAGAT TA -             #CCGCCGTT    540                                                                  - - TTTCAAGAAA TATGCACCTT CGATAGCTCC AATTGTTCCG AAAGACCAGC AG -             #GAACCACA    600                                                                  - - AACAGATTGA TCTTTAACTG GAGTAACAGC TCCATAAATT CTCCAGTCGA AA -             #GAATCTGG    660                                                                  - - TAACGGATCC TTATCTGGTA ACGGATCCTT CTTATCGTCA TCGTCGTACA GA -             #TCCCGACC    720                                                                  - - CATTTCCTGT CCACCAGTCA TGCTAGCCAT ACCATGATGA TGATGATGAT GA -             #GAACCCCG    780                                                                  - - CAT                  - #                  - #                  - #                 783                                                                   - -  - - (2) INFORMATION FOR SEQ ID NO:94:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 660 base - #pairs                                                  (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (genomic)                                      - -     (ix) FEATURE:                                                                   (A) NAME/KEY: CDS                                                              (B) LOCATION: 1..660                                                  - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:94:                               - - TTA CCA GAT TCT TTC GAC TGG AGA ATT TAT GG - #A GCT GTT ACT CCA GTT            48                                                                        Leu Pro Asp Ser Phe Asp Trp Arg Ile Tyr Gl - #y Ala Val Thr Pro Val              1               5 - #                 10 - #                 15               - - AAA GAT CAA TCT GTT TGT GGT TCC TGC TGG TC - #T TTC GGA ACA ATT GGA            96                                                                        Lys Asp Gln Ser Val Cys Gly Ser Cys Trp Se - #r Phe Gly Thr Ile Gly                         20     - #             25     - #             30                   - - GCT ATC GAA GGT GCA TAT TTC TTG AAA AAC GG - #C GGT AAT CTT GTA CGA           144                                                                        Ala Ile Glu Gly Ala Tyr Phe Leu Lys Asn Gl - #y Gly Asn Leu Val Arg                     35         - #         40         - #         45                       - - TTG TCT CAA CAG GCT TTG ATT GAT TGT TCT TG - #G GGA TAT GGA AAT AAT           192                                                                        Leu Ser Gln Gln Ala Leu Ile Asp Cys Ser Tr - #p Gly Tyr Gly Asn Asn                 50             - #     55             - #     60                           - - GGT TGT GAC GGT GGC GAG GAC TTC CGC GCT TA - #C CAA TGG ATG ATG AAA           240                                                                        Gly Cys Asp Gly Gly Glu Asp Phe Arg Ala Ty - #r Gln Trp Met Met Lys             65                 - # 70                 - # 75                 - # 80        - - CAT GGA GGA ATT CCT ACT GAA GAA GAT TAT GG - #T GGT TAC TTG GGA CAA           288                                                                        His Gly Gly Ile Pro Thr Glu Glu Asp Tyr Gl - #y Gly Tyr Leu Gly Gln                             85 - #                 90 - #                 95               - - GAT GGT TAC TGC CAT GTC AAC AAC GTT ACT TT - #A GTT GCT CCC ATC ACA           336                                                                        Asp Gly Tyr Cys His Val Asn Asn Val Thr Le - #u Val Ala Pro Ile Thr                        100      - #           105      - #           110                   - - GGG TAT GTC AAC GTA ACA CGT AAT GAC GTT GA - #C GCT ATG AAG GTT GCC           384                                                                        Gly Tyr Val Asn Val Thr Arg Asn Asp Val As - #p Ala Met Lys Val Ala                    115          - #       120          - #       125                       - - CTT CTT AAA CAC GGT CCA ATT TCG GTG GCC AT - #T GAC GCA TCA CAC AAA           432                                                                        Leu Leu Lys His Gly Pro Ile Ser Val Ala Il - #e Asp Ala Ser His Lys                130              - #   135              - #   140                           - - ACA TTC AGT TTT TAC TCC AAC GGC GTT TAC TA - #C GAA CCG AAA TGT GGC           480                                                                        Thr Phe Ser Phe Tyr Ser Asn Gly Val Tyr Ty - #r Glu Pro Lys Cys Gly            145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - AAT AAA GAG GAC GAG TTG GAC CAT GCC GTA TT - #A GTA GTC GGT TAT         GGT      528                                                                     Asn Lys Glu Asp Glu Leu Asp His Ala Val Le - #u Val Val Gly Tyr Gly                           165  - #               170  - #               175               - - GAA ATC AAC AAC GAA CCT TAC TGG TTG GTC AA - #G AAT TCC TGG TCG AAT           576                                                                        Glu Ile Asn Asn Glu Pro Tyr Trp Leu Val Ly - #s Asn Ser Trp Ser Asn                        180      - #           185      - #           190                   - - TTG TGG GGA AAT GAT GGT TAT ATT TTG ATG TC - #C GCC AGA AAT AAT AAT           624                                                                        Leu Trp Gly Asn Asp Gly Tyr Ile Leu Met Se - #r Ala Arg Asn Asn Asn                    195          - #       200          - #       205                       - - TGC GGA GTT TTG ACT GAT CCA ACT TAT GTT AC - #T ATG                      - #      660                                                                     Cys Gly Val Leu Thr Asp Pro Thr Tyr Val Th - #r Met                                210              - #   215              - #   220                           - -  - - (2) INFORMATION FOR SEQ ID NO:95:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 220 amino - #acids                                                 (B) TYPE: amino acid                                                           (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:95:                               - - Leu Pro Asp Ser Phe Asp Trp Arg Ile Tyr Gl - #y Ala Val Thr Pro         Val                                                                                1               5 - #                 10 - #                 15              - - Lys Asp Gln Ser Val Cys Gly Ser Cys Trp Se - #r Phe Gly Thr Ile Gly                    20     - #             25     - #             30                   - - Ala Ile Glu Gly Ala Tyr Phe Leu Lys Asn Gl - #y Gly Asn Leu Val Arg                35         - #         40         - #         45                       - - Leu Ser Gln Gln Ala Leu Ile Asp Cys Ser Tr - #p Gly Tyr Gly Asn Asn            50             - #     55             - #     60                           - - Gly Cys Asp Gly Gly Glu Asp Phe Arg Ala Ty - #r Gln Trp Met Met Lys        65                 - # 70                 - # 75                 - # 80        - - His Gly Gly Ile Pro Thr Glu Glu Asp Tyr Gl - #y Gly Tyr Leu Gly Gln                        85 - #                 90 - #                 95               - - Asp Gly Tyr Cys His Val Asn Asn Val Thr Le - #u Val Ala Pro Ile Thr                   100      - #           105      - #           110                   - - Gly Tyr Val Asn Val Thr Arg Asn Asp Val As - #p Ala Met Lys Val Ala               115          - #       120          - #       125                       - - Leu Leu Lys His Gly Pro Ile Ser Val Ala Il - #e Asp Ala Ser His Lys           130              - #   135              - #   140                           - - Thr Phe Ser Phe Tyr Ser Asn Gly Val Tyr Ty - #r Glu Pro Lys Cys Gly       145                 1 - #50                 1 - #55                 1 -       #60                                                                               - - Asn Lys Glu Asp Glu Leu Asp His Ala Val Le - #u Val Val Gly Tyr         Gly                                                                                              165  - #               170  - #               175              - - Glu Ile Asn Asn Glu Pro Tyr Trp Leu Val Ly - #s Asn Ser Trp Ser Asn                   180      - #           185      - #           190                   - - Leu Trp Gly Asn Asp Gly Tyr Ile Leu Met Se - #r Ala Arg Asn Asn Asn               195          - #       200          - #       205                       - - Cys Gly Val Leu Thr Asp Pro Thr Tyr Val Th - #r Met                           210              - #   215              - #   220                           - -  - - (2) INFORMATION FOR SEQ ID NO:96:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 25 amino - #acids                                                  (B) TYPE: amino acid                                                           (C) STRANDEDNESS:                                                              (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: protein                                            - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:96:                               - - Xaa Pro Pro Pro Glu Met Leu Gly Gly Pro Se - #r Ile Phe Ile Phe Pro       1               5   - #                10  - #                15                - - Pro Lys Lys Lys Asp Asp Leu Leu Ile                                                   20      - #            25                                           - -  - - (2) INFORMATION FOR SEQ ID NO:97:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 25 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:97:                               - - TTGGGATACA CTTTGACTGT TAACC          - #                  - #                    25                                                                       - -  - - (2) INFORMATION FOR SEQ ID NO:98:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 24 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:98:                               - - GTGAGCAACC ATTATTTCCA TATC          - #                  - #                     24                                                                       - -  - - (2) INFORMATION FOR SEQ ID NO:99:                                     - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 23 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:99:                               - - TGGGTWGTWA CWGCWGCWCA TTG           - #                  - #                     23                                                                       - -  - - (2) INFORMATION FOR SEQ ID NO:100:                                    - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 22 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:100:                              - - ATTCCTCGTG GTTCAGTCGC TC           - #                  - #                      22                                                                       - -  - - (2) INFORMATION FOR SEQ ID NO:101:                                    - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 26 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:101:                              - - GAAGATGTWG ATATTTCWAC ATGTGG          - #                  - #                   26                                                                       - -  - - (2) INFORMATION FOR SEQ ID NO:102:                                    - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 27 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:102:                              - - GAAAATGAAA TCCACTTAAA CATTACG          - #                  - #                  27                                                                       - -  - - (2) INFORMATION FOR SEQ ID NO:103:                                    - -      (i) SEQUENCE CHARACTERISTICS:                                                  (A) LENGTH: 21 base - #pairs                                                   (B) TYPE: nucleic acid                                                         (C) STRANDEDNESS: single                                                       (D) TOPOLOGY: linear                                                  - -     (ii) MOLECULE TYPE: DNA (primer)                                       - -     (xi) SEQUENCE DESCRIPTION: SEQ ID NO:103:                              - - CTCTTATTGT ACGAGGGATG C           - #                  - #                       - #21                                                                  __________________________________________________________________________

While various embodiments of the present invention have been described in detail, it is apparent that modifications and adaptations of those embodiments will occur to those skilled in the art. It is to be expressly understood, however, that such modifications and adaptations are within the scope of the present invention, as set forth in the following claims. 

What is claimed is:
 1. An isolated protein selected from the group consisting of: (a) a protein comprising an amino acid sequence selected from the group consisting of SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:16, SEQ ID NO:19, SEQ ID NO:22, SEQ ID NO:24, SEQ ID NO:27, SEQ ID NO:30, SEQ ID NO:33, SEQ ID NO:36, SEQ ID NO:38, SEQ ID NO:41, SEQ ID NO:44, SEQ ID NO:67, SEQ ID NO:68, SEQ ID NO:69, SEQ ID NO:70, SEQ ID NO:71 and SEQ ID NO:72; and (b) a flea protein comprising a homologue of said protein of (a), wherein daid homologue comprises at least 6 contiguous amino acids of an amino acid sequence selected from the group consisting of SEQ ID NO:27, SEQ ID NO:30, SEQ ID NO:33, SEQ ID NO:36, SEQ ID NO:38, SEQ ID NO:41, SEQ ID NO:44, SEQ ID NO:67, SEQ ID NO:68, SEQ ID NO:71 and SEQ ID NO:72.
 2. The protein of claim 1, wherein said protein comprises an amino acid sequence selected from the group consisting of SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO: 16, SEQ ID NO: 19, SEQ ID NO:22, SEQ ID NO:24, SEQ ID NO:27, SEQ ID NO:30, SEQ ID NO:33, SEQ ID NO:36, SEQ ID NO:38, SEQ ID NO:41, SEQ ID NO:44, SEQ ID NO:67, SEQ ID NO:68, SEQ ID NO:69, SEQ ID NO:70, SEQ ID NO:71 and SEQ ID NO:72.
 3. The protein of claim 1, wherein said protein is encoded by a nucleic acid molecule comprising a nucleic acid sequence selected from the group consisting of: SEQ ID NO:9, SEQ ID NO:12, SEQ ID NO:15, SEQ ID NO:18, SEQ ID NO:21, SEQ ID NO:23, SEQ ID NO:26, SEQ ID NO:29, SEQ ID NO:32, SEQ ID NO:35, SEQ ID NO:37, SEQ ID NO:40, SEQ ID NO:43 and SEQ ID NO:66. 